BIOADHESIVES

Abstract: Vectors that are capable of expressing, in microbial transformants, a protein having the native amino acid sequence of a bioadhesive precursor protein of a marine animal selected from the group consisting of mussels, barnacles, and oysters. The bioadhesive precursor protein can be expressed in transformants, recovered and converted to a bioadhesive protein by hydroxylation.

Description: -/- BIOADHESIVES CROSS-REFERENCE TO RELATED APPLICATIONS This application is a continuation-in-part of application Serial No. 933,945, filed November 24, 1986, which is a continuation-in-part of ap¬ plication Serial No. 650,128, filed September 13, 1984. BACKGROUND OF THE INVENTION Field of the Invention This invention relates to the production of bio- adhesives that can be employed to bond substances for use in wet environments. Typically, the bioadhesives of the invention are employed as marine adhesives, biomedical adhesives or dental adhesives. The inven¬ tion further relates to the microbial production of bioadhesive precursor proteins that can be converted to bioadhesives by chemical or enzymatic treatment. Brief Description of Background Art The properties of adhesives generally must be tailored to meet the requirements of the particular environments in which they are to be used. Ideally, an adhesive should be cured and it should maintain both its adhesivity and cohesivity under the condi¬ tions of use. Curing is the altering of the physical properties of an adhesive by chemical or enzymatic means. In the case of the bioadhesives produced by the procedures described herein, curing is likely to be due to the cross-linking of adjacent uncured adhesive molecules by catalytic and/or chemical agents. Curing may also involve adhesive cross-linking with the substrate. Many adhesives that exhibit excellent adhesive properties under dry conditions suffer a substantial or total loss of those properties in wet environments. Furthermore, adhesives of the prior art cannot be cured in vet environments. Consequently, it has been particularly difficult to develop adhesives for use in wet environments, such as marine adhesives or adhe¬ sives for use in medical and dental applications. Marine mussels and other sessile invertebrates have the ability to secrete adhesive substances by which they affix themselves to underwater objects. For example, mussels of the genus Mytilus, e.g., the species Mytilus edulis and Mytilus californianus, de¬ posit an adhesive substance from the mussel foot that becomes cured, forming a permanent attachment to the substrate. A major component of the adhesive plaque deposited by __ edulis has been identified as a hy- droxylated protein of about 130,000 daltons ( aite, J. H., J. Biol. Chem. , 258_:2911-2915 (1983)). While this substance might make an excellent adhesive for use in wet environments, isolation of the uncured adhesive from mussels for commercial use is not practical, since the extraction of 1 kg of the adhesive substance would require the use of about 5 to 10 million mussels. Besides involving a very laborious process, great care would have to be taken, in case the adhe¬ sive were to be employed in medical applications, to insure that other mussel proteins and contaminants were removed from the bioadhesive prior to use in or¬ der to prevent antigenic or anaphylactic reactions. U.S. Patent No. 4,585,585 describes a procedure for preparing a bioadhesive polymer by chemically linking decapeptide units produced by the enzymatic digestion of isolated mussel adhesive protein. In accordance with the disclosure of that patent, a bio¬ adhesive protein \s first isolated from phenol glands of mussels of the genus Mytilus using the protein purification procedures described by Waite and Tanzer in Science, 212:1038 (1981). The isolated bioad¬ hesive, having a molecular weight of 120,000 to 140,000 daltons, is first treated with collagenase, which reduces its molecular weight by about 10,000 daltons. The treated protein is then digested with trypsin, and the digested protein subjected to gel filtration dialysis to isolate decapeptides of the general formula NH 2 -Ala-Lys-Pro/Hyp-Ser/Thr-Tyr/Dopa-Pro/Hyp- J?ro/Hyp-Ser/Thr-Tyr/Dopa-Lys-COOH The decapeptides produced in this manner are then polymerized by the use of-chemical linking groups such as glutaraldehyde,. oligopeptides, amino acids or other, bifunctiόnal linking groups to produce bioadhesives containing up to about 1,000 such decapeptide units. The procedure of U.S. Patent No. 4,585,585 still requires the isolation of bioadhesive protein from mussel, which, as previously indicated, is impracti¬ cal on a commercial scale. Moreover, in addition to the laborious., purification procedure, this process adds the additional steps of enzymatic digestion, iso¬ lation of the decapeptide fragments and chemical reas- semblage of the fragments into a bioadhesive polymer. This arduous procedure is not well-suited to commer¬ cial production. Further, the polymers produced by this method are quite different from the mussel ad¬ hesive since they contain only chemically polymerized decapeptides produced by trypsin digestion of the natural molecule. Analysis of the natural gene described herein demonstrates that there are other significant sequence elements in the mussel adhesive from Mytilus edulis. Thus a need has continued to exist for means and methods for the efficient production of bioadhesives having the excellent properties associated with the mussel foot bioadhesive in wet environments. A further need has continued to exist for means and methods for producing bioadhesives having the pro¬ perties of the mussel foot adhesive without the neces¬ sity of handling and processing large quantities of mussels. SUMMARY OF THE INVENTION This invention involves application of the tech¬ niques of recombinant DNA technology to the production of bioadhesives of the type produced by marine animals such as mussels, barnacles and oysters. In particu¬ lar, we have cloned genes for native bioadhesive pre¬ cursor proteins (also termed polyphenolic proteins) of the mussel Mytilus edulis and constructed expression vectors capable of expressing these precursor proteins in microbial hosts. The bioadhesive precursor proteins are chemically or enzymatically hydroxylated, mimicking a process which occurs ijn vivo in the mussel, to produce a bioadhesive protein which cures to produce an adhesive having excellent properties in wet environments. In the course of our work, we have isolated multi¬ ple cDNA clones coding for native M. edulis bioad¬ hesive precursor proteins. Two of the encoded pro- teins (from cDNA isolates 14-1 and 52) have a common C-terminal sequence of 134 amino acids, including a hexapeptide followed by twelve decapeptides, ten of which display non-homology with the decapeptide sequence described in U.S. Patent No. 4,585,585. Another encoded protein, from cDNA isolate 55, has homology with 53 and 110 amino acids in the proteins encoded by cDNAs 14-1 and 52, as well as homology in a different region with 34 and 57 amino acids of the proteins encoded by cDNA isolates 56 and 52. The cod¬ ing homologies are conserved at the DNA level. One likely explanation for these observations is that the M. edulis polyphenolic protein gene consists of sever¬ al exons that can be spliced together in different combinations at the mRNA level. Alternatively, there could be several closely related. genes. This disclosure is not intended to be limiting to any one exon combination or bioadhesive - precursor protein gene, as a method is described which allows the isolation and use of any existing- gene or gene_ combinations. In all the unique decapeptide coding segments observed in the 4 cDNA clones described herein, 24 of 39 decapeptides display non-homolog with the decapeptide described in U.S. Patent 4,585,585. Yet a further encoded protein, from cDNA isolate Nl, a 2.1 kb clone, contains 76 continuous tandem repeats of decapeptides and hexapeptides. Of the 63 decapeptides in the Nl protein, 32 display non-homology with the decapeptide described in U.S. Patent 4,585,585. There is provided, in accordance with this inven¬ tion, a DNA segment comprising codons for the native amino acid sequence of a bioadhesive precursor protein of a marine animal, e.g., of a mussel of the genus My¬ tilus, e.g., M. edulis. In another embodiment of the invention, vectors are provided for expression of bioadhesive precursor protein comprising: (a) a DNA sequence comprising codons for the native amino acid sequence of a bioadhe¬ sive precursor protein of a marine ani¬ mal, e.g., of a mussel, e.g., a mussel of the genus Mytilus, or fragment there¬ of; and (b) a promoter and transcription initiation signal, operably linked to said DNA seg¬ ment, which are capable of effecting microbial expression of the bioadhesive precursor protein. In yet another embodiment of the invention, there are provided transformant microorganisms capable of expressing bioadhesive precursor proteins that can be chemically or enzymatically converted to a bioadhe¬ sive. The transformants are produced by transforming a host microorganism such as an E. coli or Saccharo- myces cerevisiae with the aforementioned expression vector. In another embodiment of the invention, there is provided a method of producing a bioadhesive protein. A transformant microorganism containing the expression vector for the bioadhesive precursor protein is cul¬ tured and subjected to conditions under which the pre¬ cursor protein is expressed and the precursor protein is recovered. The precursor protein, as will be de¬ scribed in detail below, contains amino acid sequence subunits which possess tyrosine and proline residues. The precursor protein is converted into a bioadhesive protein by chemical or enzymatic hydroxylation, which converts at least a portion of the tyrosine residues to 3,4-dihydroxyphenylalanine (DOPA) residues and, optionally, at least a portion of the proline residues to hydroxyproline residues. In a preferred embodiment of the invention, the bioadhesive precursor protein is hydroxylated by enzymatic treatment, for example, with mushroom tyrosinase. There are also provided by the invention methods of bonding surfaces which involve application of the bioadhesive protein, either as an adhesive per se or as a primer coating for an adhesive, to the surfaces being bonded. There are also provided by the invention methods for bonding or wetting surfaces with composite materi¬ als including bioadhesive protein and other polymers and/or adhesives. BRIEF DESCRIPTION OF THE DRAWINGS FIG. 1 presents he DNA sequence and translated amino acid sequence of a gene identified as cDNA clone . 14-1, which codes for a bioadhesive precursor protein of M. edulis. FIG. 2 presents the DNA sequence and translated amino acid sequence of a gene identified as cDNA clone 52, which codes for a bioadhesive precursor protein of M. edulis. FIG. 3 presents the DNA sequence and translated amino acid sequence of a gene identified as cDNA clone 55, which codes for a bioadhesive precursor protein of M. edulis. FIG. 4 presents the DNA sequence and translated amino acid sequence of a gene identified as cDNA clone 56, which codes for a bioadhesive precursor protein of M. edulis. FIG. 5(a) presents the sequence of a portion of a multi-restriction site region of plasmid pGX2627. Fig. 5(b) presents a DNA sequence and translated amino acid sequence of portions of a synthetic DNA segment employed in the construction of plasmid pGX2346 from plasmid pGX2287. FIG. 6 is a schematic representation of the con¬ struction of plasmids pGX2346 pGX2368, pGX2377, pGX2380, pGX2381 and ρGX2383. Plasmid pGX2383 is an expression vector for a bioadhesive precursor protein. FIG. 7 is the DNA sequence of plasmid pGX2627. FIG. 8 is a restriction map of the YpGX265GAL4 shuttle vector showing the position and orientation of the major functional elements, including the GAL4 binding site, the MF-alpha 1 transcription initiation site, the PH05 signal encoding sequence, the GAPDH terminator sequence, the E. coli ampicillin-resistance marker and the LEU2 marker for plasmid selection in S_. cerevisiae. Figure 9 presents the DNA sequence and translated amino acid sequence of a gene identified as cDNA clone Nl, which codes for a bioadhesive precursor protein of M. edulis. Figure 10 presents the amino acid variations in decapeptides of clone Nl. The variations in each position are shown, with the subscripts indicating the number of decapeptides in clone Nl in which the particular amino acid occurs at the position cited. Figure 11 is a restriction map of YpGX283, showing the position and orientation of the major functional elements. Figure 12 is a restriction map of plasmid YpGX405 yeast ^ coli shuttle vector, showing the position and orientation of major functional elements. -9- DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS The preparation of an expression vector of the invention generally involved the following series of steps: (a) Isolating RNA from the marine animal, e.g., from the phenol gland of the foot of M. edulis; (b) Synthesizing complementary DNA (cDNA) and inserting the cDNA into bacteriophage; (c) Transfecting E. coli with the recombinant bacteriophage and culturing the transfected E. coli? (d) Selecting clones containing bioadhesive pre¬ cursor protein genes by hybridization of transformants with a radiolabeled probe; (e) Insertion of a DNA segment coding for a bio¬ adhesive precursor protein into an expression vector as the 3* portion of a gene coding for a hybrid fusion protein. In some cases, a methionine residue is located on the amino. terminal . side of the amino acid sequence of the bioadhesive precursor protein. (By the term "DNA segment" is intended a portion, but less than the entirety, of the genome of the marine animal. ) The mRNA can be isolated from M. edulis by isolat¬ ing the foot organ containing the phenol gland from live mussels; homogenizing the tissue in liquid nitro¬ gen; dissolving the frozen tissue in guanidine thio- cyanate as described by McCandliss, R. et a_l. , Methods Enzymol. y 7JJ:51-57 (1981), or lysing the frozen tissue in the presence of ribonucleoside-vanadyl complexes and extracting the lysed tissue with phenol as described by Berger, S. et al.., J. Biol. Chem. , 255:2955-2961 (1980); and purifying poly A-containing RNA by binding on oligo-dT cellulose as described by Aviv, H. and Leder, P., Proc. Nat . Acad. Sci. USA, 69 1408-1412 (1972). Similar procedures may be used in isolating the mRNA transcript of the gene of other marine animals encoding bioadhesive protein. The mRNA extracted, i.e., from the phenol glands, is then employed as a template to produce cDNA in the presence of reverse transcriptase, dATP, dCTP, dGTP and dTTP, according to well-known procedures. Advan¬ tageously, a radioactive marker such as [alpha- 32PldCTP is used to monitor the synthesis. The mRNA moiety is then removed using known procedures and a second strand of cDNA is synthesized, using the -first strand as a template, in the presence of E. coli DNA polymerase I (Klenow fragment), dATP, dCTP, dGTP and dTTP. As those skilled in the art are aware, the double-stranded (ds-cDNA) DNA produced in this manner contains a hairpin loop. The single-stranded loop can conveniently be removed by digestion with SI nuclease. The ds-cDNA produced in the manner described is then inserted into a cloning vector in order to pro¬ duce a library of clones to screen for the presence of the bioadhesive precursor protein gene. We employed the- bacteriophage lambda gtlO as a cloning vector; however, those skilled in the art will be aware that any of numerous other vectors could conveniently be employed to the same end. In order to insert the ds- cDNA into the EcoRI site of lambda gtlO, we first added synthetic 8-base pair EcoRI linkers to both ends of the ds-cDNA after blunting the ends of the ds-cDNA in the presence of DNA polymerase I (Klenow fragment) and dATP, dCTP, dGTP, and dTTP. To prevent cleavage of the ds-cDNA by EcoRI it was first methylated with -11- EcoRI methylase. The linkers were phosphorylated in polynucleotide kinase buffer and ligated to the ends of the blunted ds-cDNA in the presence of T4 ligase. Multiple linkers were then removed from the ends by digestion with EcoRI. The ds-cDNA with EcoRI linkers was inserted into EcoRI-cut lambda gtlO. The cloning vector containing the cDNA is propa¬ gated in a suitable host. In the case of the recom- binant lambda gtlO, we packaged the recombinant phage DNA into bacteriophage lambda heads for introduction into E. coli strain BNN102 (Huynh, T.V. et ad.. Con¬ structing and screening cDNA libraries in lambda gtlO and lambda gtll, I_n:D.M. Glover (ed.) DNA cloning. Vol. I, IRL Press, Oxford (1985), pp. 49-78) by pro¬ cedures described by Enquist and Sternberg, Methods Enzymol., 6J 281-298 (1979). On this host strain, only lambda gtlO bacteriophage-carrying inserts at the EcoRI site will form plaques. The transfected hosts are cultured under appropriate conditions and examined, for plaques. The ds-cDNA library produced in. the above manner is screened for the presence of the bioadhesive pre¬ cursor protein gene by hybridization with a radio- labeled synthetic oligonucleotide probe. One pre¬ dominant decapeptide in the bioadhesive precursor protein is known to be ala-lys-pro-ser-tyr-pro-pro- thr-tyr-lys. To screen the recombinant phage library for the precursor protein gene, we constructed and radiolabeled two 30-base synthetic oligonucleotide probes coding for the decapeptide, these two selected randomly from the 98,304 possible coding combinations. The two sequences selected were S87/03048 GCG AAA CCA AGT TAC CCA CCG ACC TAC AAA and GCG AAA CCT TCT TAT CCG CCT ACC TAT AAG. Using hybridization washing conditions with de¬ creased stringency that were previously established in Southern blot experiments and which are described in detail in the examples which follow, the radiolabeled probes were used to screen plaques which had been fixed on duplicate nitrocellulose filter replicates. To our knowledge, the 30-base probes are the shortest synthetic probes that have been used successfully in this type of screening. Plaques that gave positive signals by autoradiography of duplicate filters were purified, screened again and grown as plate lysates for DNA preparation as described in Maniatis, T. e_t al. , "Molecular Cloning: A Laboratory Manual," Cold Spring Harbor Laboratory, pp. 371-372 (1982). The DNA sequence of the isolated clones can be determined by well-known methods. Using the procedures described generally above and described in greater detail in the examples that fol¬ low, we isolated several clones. Five have been char¬ acterized in detail and are identified herein as clones 14-1, 52, 55, 56, and Nl, which contained DNA sequences coding for bioadhesive precursor proteins. Fig. 1 presents the DNA sequence and the translated amino acid sequence of the insert in clone 14-1. The DNA segment comprising the insert in clone 14-1 con¬ tains codons for a 203-amino acid sequence. In Fig. 1, the sequence has been divided to show 20 subunits, including 19 decapeptides and 1 hexapeptide (subunit no. 8). Fig. 2 presents the DNA sequence and the trans¬ lated amino acid sequence from the insert in cDNA -13- clone 52. This DNA segment in the insert in cDNA clone 52 contains codons for a 334-amino acid sequence. In Fig. 2, the sequence has been divided to show 26 subunits which include 23 decapeptides and 3 hexapeptides, and an amino-terminal proline-rich seg¬ ment. This cDNA clone and two of the other analyzed clones (55, 56) contain some noncoding DNA sequence (not shown) which we believe is derived from intron sequences. This suggests that some of the clones were derived from incompletely processed nuclear RNA. The DNA sequences of the cDNA clones 14-1 and 52 display extensive homology at their 3'-ends. In par¬ ticular, the last 138 codons of the translated regions are identical and include codons for a hexapeptide followed by 12 decapeptides (beginning at codon 75 of clone 14-1 in Figure 1 and codon 205 of clone 52 in Figure 2). Clone 52 contains a segment at the 5'-end that codes for a proline-rich segment of 82 amino acids. There is also a direct repeat of DNA sequences in clone 52. Specifically, the DNA-for codons 92-147 is exactly repeated in codons 148-203. DNA sequences for cDNA clones 55 and 56 are shown in Figures 3 and 4. Codons 19-129 of cDNA clone 55 match the codons 148-259 of cDNA clone 52 with one exception. Codon 65 of clone 55 is an alanine while the clone 52 corresponding codon 195 is isoleucine. There is also homology between the cDNA of clone 55 (Fig. 3) and the cDNA of clone 56 (Fig. 4). The codons 1-33 of clone 55 match codons 54-86 of clone 56. Some of the sequences are in the area where clone 55 also overlaps with clone 52, and in the direct repeating segment of clone 52. The DNA sequence for clone Nl is shown in Figure 9. Clone Nl does not have a perfect overlap with any of clones 14-1, 52, 55 or 56. Clone Nl also does not have a characteristic 5'-end sequence and therefore does not, in all likelihood, include the N-terminus of the natural gene. It does, however, code for a poly- peptide with a molecular weight of about 80,000, a substantial portion of the natural polypeptide (mole¬ cule weight of 110,000-130,000). Thus it can be seen that the cDNA clones are related in a complex way. Although there are areas in common, there is also generally divergence, suggesting either the existence of multiple genes or varied patterns of mRNA splicing for one gene or both. As previously indicated, this invention provides expression vectors for microbial expression of bio¬ adhesive precursor proteins. The expression vectors contain a DNA segment coding for a bioadhesive precur¬ sor protein such as that of a mussel of the genus My¬ tilus or a fragment thereof, the DNA segment being operably linked to a promoter and transcription initi¬ ation signals that are capable of effecting microbial expression. In one embodiment, the DNA segment con¬ tains codons for at least the portion of the amino acid sequence comprising subunits 8-20 of clone 14-1 and subunits 14-21 of clone 52; that is, amino acids 72-197 of clone 14-1 and amino acids 201-326 of clone 52. Figures 6 and 7 outline one procedure that is employed to produce expression vectors of the inven¬ tion. These expression vectors are capable of effect¬ ing expression of a bioadhesive precursor protein as a hybrid fusion protein which, in some cases, contains a methionine residue just upstream of the protein of interest. Since the bioadhesive precursor proteins encoded by all of the clones described above except clone Nl contain no methionine residues, they can be conveniently separated from the fused sequences by treatment with cyanogen bromide, which cleaves the fusion protein at methionine residues. As regards the protein encoded by the Nl clone, cyanogen bromide treatment may cleave at the methionine residue found in the second decapeptide, still leaving essentially the entire polypeptide intact.As the amino-terminal non-bioadhesive precursor protein amino acids will comprise at most roughly 15%, and in many cases much less, of the total adhesive protein precursor, removal of those amino acids by cyanogen bromide cleavage will not be necessary for all applications. Referring to Fig. 6, the DNA segment containing the coding region for the bioadhesive precursor pro¬ tein of clones 14-1 and 52 was obtained from the lambda gtlO cloning vectors by EcoRI digestion and. subcloned into the EcoRI site of plasmid pGX2627. The resultant plasmids were designated pGX2368 and PGX2377. Plasmids pGX2368 and pGX2377 were subjected to oligonucleotide-directed mutagenesis (Zoller, M.J. et al. , Methods Enzymol. , Vol 100B, R. Woo, L. Grassa, and K. Moldave (eds.), Academic Press, New York, pp. 468-500 (1983)) to create Bell sites located at the translation terminators of each gene (see Figs. 1 and 2). Since pGX2627 is a derivative of pGXl066 (ATCC No. 39955) that contains an M13 origin of replication, the pGX2368 and pGX2377 derivatives of pGX2627 were isolated in a single-stranded form for the oligonucle¬ otide-directed mutagenesis experiments by infection of cultures with IRl or M13 bacteriophage. The new plasmids containing Bell sites were designated pGX2380 and pGX2381. The DNA segments containing the coding regions for the bioadhesive precursor proteins were then inserted into E. coli and yeast expression vectors in the -16- proper reading frame. In an E. coli example presented below, the cDNA from phage 14-1 located in pGX2380 was cloned as a trpB gene fusion starting with an expres¬ sion vector identified as pGX2287 that was designed for bovine chymosin production. Construction of this vector is described in detail in copending, commonly assigned U.S. Application Serial No. 671,976. Plasmid pGX2287 has been deposited with the USDA Northern Re¬ gional Research Laboratory, Peoria, Illinois, with ac¬ cession No. NRRL-B15788. It contains the trpB bovine chymosin gene under the control of the leftward opera¬ tor and rightward promoter regions of phage lambda (see copending, commonly assigned U.S. application Serial No. 534,982) and an appropriately positioned transcription initiation signal. It also contains an ampicillin-resistance gene. Plasmid pGX2287 was cut with Clal, which cleaved the plasmid in trpB 24-base pairs upstream from the 5'-end of the chymosin coding sequence. A 116-base pair Clal-Sphl synthetic DNA was then ligated with linearized plasmid. The sequence of the synthetic DNA fragment which was ligated to the linearized plasmid is given in Fig. 5(b). The ligated DNA was then di¬ gested with S I and the large fragment was isolated, resulting in the removal of a substantial portion of the chymosin coding region. The plasmid was then re- circularized with T4 ligase. The resultant plasmid, identified pGX2346, contained the synthetic DNA frag¬ ment in place of the central portion of the chymosin gene. Other specific vector constructions for the ex¬ pression of the bioadhesive precursor protein will be apparent to those skilled in the art based on the de¬ scription herein. As a general rule, however, it is advantageous to construct the vector by inserting the bioadhesive precursor coding region as an in-frame fusion with another gene that is under the control of an efficient promoter. Preferably, the fusion is con¬ structed such that the encoded fusion protein contains a methionine residue just upstream, e.g., within about 10 residues, from the bioadhesive precursor protein segment. In the specific construction described here¬ in, pGX2383, there is a methionine residue 7 amino acids upstream from the bioadhesive precursor sequence. The recovered bioadhesive precursor protein can be treated with cyanogen bromide, using conditions well-known in the art, to remove extraneous amino acid sequences. As those skilled in the art are aware, cyanogen bromide cleaves proteins at methionine resi¬ dues. Since there are no internal methionine resi¬ dues within the bioadhesive precursor protein itself, the protein remains intact. It will be readily apparent that modifications can be made in the vector construction such that the ex¬ pression vector can carry the entire coding region for the bioadhesive precursor protein or a coding region for a fragment thereof. Preferably, the fragment con¬ tains at least about 5-20% of the coding sequence of a native bioadhesive precursor protein of a marine ani¬ mal, e.g., of a mussel, e.g., of a mussel of the genus Mytilus. Encoded amino acids fused to the N-terminal or C-terminal ends of the encoded bioadhesive precur¬ sor, which cannot be removed by cyanogen bromide cleavage, are not believed to interfere with the func¬ tioning of the bioadhesive protein. The expression vector provided by the invention is used to transform any suitable host microorganism, using known means, to produce a transformant. Suit¬ able host organisms include, for example, E. coli or other related gram-negative organisms such as Salmo- nella, Klebsiella, Erwinia, etc. With plasmids related to pGX2383 the host preferably contains the lambda cI857 gene coding for temperature-sensitive lambda represser. In the case of pGX2383, we chose to employ E. coli strain GX3Q1S as the host. This was because pGX2383 contains the trpED portion deleted in the trpEDl02 chromosomal deletion in GX3015. The plasmid is therefore stabilized by growing the trans¬ formant in a tryptophan-deficient medium. In a different embodiment, an expression vector may be used to transform yeast, e.g., Saccharomyces cerevisiae. Typical vectors of this type are dis¬ closed in U.S. application Serial No. 918,147, filed on October 14, 1986, and having the title "Composite Yeast Vectors," incorporated by reference herein. One preferred vector for expression of bioadhesive pre¬ cursor protein in yeast comprises the yeast shuttle vector YpGX265GAL4 (ATCC #67233). This vector is characterized by a promoter that is a hybrid-derived from the S. cerevisiae GAL1 and MF-alpha 1(alpha-factor) promoters. This promoter system permits galactose-regulated expression. The regula¬ tory gene comprises the GAL4 gene which encodes GAL4 protein, a positive regulator of the GALl-MF-alpha 1 hybrid promoter derived from §. cerevisiae. The terminator in the YpGX265GAL4 vector system is derived from synthetic DNA and is based on S. cerevisiae GAPDH transcription terminator. The signal encoding sequence, also derived from synthetic DNA, is based on the S_. cerevisiae PH05 signal. Codons are designed substantially for usage preference in S. cerevisiae. The YpGX265GAL4 vector contains the LEU2 gene, a marker for plasmid selection in S. cerevisiae. It also contains DNA derived from S. cerevisiae 2-micron plasmid which provides a plasmid replication origin for S. cerevisiae. The vector is further charac¬ terized by the E. coli replication origin derived from pAT153, and an E. coli selectable marker which is am icillin resistance, also derived from ρAT153. The heterologous gene (the gene coding for bioadhesive precursor protein) is inserted between the PH05 signal-encoding sequence and the GAPDH terminator. A typical shuttle vector preparation is as fol¬ lows. YpGX265GAL4 is digested with restriction endo- nuclease Hindlll and large and small DNA fragments are gel-purified. The larger fragment is digested with restriction endonuclease Smal, generating two DNA fragments. M13mp9 (commercially available) is di¬ gested with Smal and Hindlll and treated with calf al¬ kaline phosphatase. The DNA fragments created by the Smal digestion of the larger Hindlll fragment of YpGX265GAL4 and from digestion of the M13mp9 are ligated and used for transformation of E. coli. The transformants containing phage double-stranded DNA with the yeast promoter-terminator cassette are identified, digested with restriction endonucleases (EcoRV and BamHI ) and the large fragment (approximate¬ ly 8 kilobase) is gel-purified. A plasmid containing bioadhesive precursor protein cDNA (i.e., pGX2380) is digested with restriction endonuclease Xbal, and the single-stranded overhang is filled with the Klenow fragment of DNA polymerase. The DNA molecule result¬ ing from the Xbal and DNA polymerase treatment of pGX2380 is digested with Bell. The resulting approxi¬ mately 625-bp DNA fragment is gel-purified. Oligonu¬ cleotide linker sequences are annealed and used in a three-way ligation with the DNA fragments created by restriction of the phage double-stranded DNA and the -20- cDNA-derived DNA molecule with the modified Xbal and Bell end. E. coli transformants containing the desired phage DNA molecules are identified by restriction endonucle¬ ase digestion. This phage DNA may be digested with restriction endonucleases (Smal and Hindlll), with the small (approximately 1,300 base pair) fragment gel-purified. YpGX265GAL4 is digested with Smal and Hindlll and the large fragment is gel-purified. The DNA molecules created by the Smal-HindiII digestions are ligated and used to transform E. coli. The desired plasmid may be linearized with restriction endonuclease (Hindlll) and treated with calf alkaline phosphatase. The 3.65-kb GAL4-containing fragment obtained from the original restriction digestion may by ligated with the linearized DNA and used to trans¬ form E. coli. Saccharomyces strains carrying mutations in the LEU2 structural gene (e.g., AH22 (ATCC # 38626)) may be transformed with this plasmid, utilizing standard methods. The yeast strain may be grown in an appro¬ priate medium (YNBD, containing 0.7% yeast nitrogen base, 2% glucose, 20 mg/liter L-histidine) to maintain the plasmid. While the above typifies one construction tech¬ nique for yeast expression, it is readily apparent that one with ordinary skill can impart modifications and variations within the general teaching. As with the vectors described above, the expression vector may comprise the entire coding region for the bioadhesive precursor protein or coding regions for fragments thereof. For production of the bioadhesive precursor pro¬ tein, the transformed yeast strain may be grown in an -21- appropriate medium. One suitable medium contains 1% yeast extract, 2% peptone, 1% glucose, and 1% galac- tose. The transformant microorganism is cultured under conditions suitable for growth and expression of the bioadhesive precursor protein gene. After the protein has been expressed, it is recovered from the trans¬ formant by known methods such as by mechanical or chemical lysis of the cells. The protein can be puri¬ fied using procedures known in the art, including well-known chromatographic procedures. The bioadhe¬ sive precursor protein is preferably purified to homo¬ geneity or near homogeneity. In the case of a fusion protein such as that expressed by pGX2383, the recovered protein can be subjected to cyanogen bromide cleavage to remove extraneous peptide sequences. The recovered bioadhesive precursor protein is converted to a bioadhesive by hydroxylation. In par¬ ticular, it is likely to be necessary to hydroxylate at least a portion of the tyrosine or tyrosine and proline residues, an event that occurs i_n vivo in the mussel. Hydroxylation converts tyrosine residues to DOPA residues and, optionally, proline residues to hydroxy proline residues. The DOPA hydroxyl groups are believed to displace water at the bond surfaces, thus contributing to the excellent wet strength of the adhesive, and DOPA residues oxidized to quinones participate in intermolecular cross-linking which cures the adhesive and imparts cohesivity. Any suitable chemical or enzymatic means for ef¬ fecting hydroxylation can be employed. It is pre¬ ferred, however, to effect hydroxylation enzymatically using an enzyme such as mushroom tyrosinase or Strep- to yces antibioticus tyrosinase. Enzymatic hydroxyla- tion procedures using these enzymes are carried out as generally described by Ito et al. , Biochem. J. 222: 407-411 (1984) and Marumo and Waite, Biochem. Biophys. Acta 872:98-103 (1986). Preferably, at least about 10% of the tyrosine residues are hydroxylated. The mushroom tyrosinase is removed from the protein using known procedures such as binding to a LH-Sephadex 60 column followed by elution with 0.2 M acetic acid or by membrane filtration. The bioadhesive protein can be lyophilized for reconstitution as an adhesive formulation at a later date. The bioadhesive protein can be employed in the form of a solution in a suitable solvent with or with¬ out other adhesive substances. Suitable solvents for the bioadhesive-include water or aqueous solutions of alcohols such as methanol, ethanol, propanol, and the like, acetone, DMSO, dimethyl formamide, and the like. The concentration of bioadhesive protein in the solution can range from very low to very high, depending upon the intended application. The bioadhesive protein or -a- solution or formula¬ tion containing the bioadhesive protein can be employ¬ ed as a primer, i.e., a preadhesive film or coating to improve the total adhesive bond to a surface. It may also be employed as a component in an * adhesive system to impart or improve water-resistant adhesivity. Another use within the scope of the invention includes use as a thin film membrane, or as a component in a thin film or membrane, particularly to impart desir¬ able permeability characteristics and/or moisture resistance to such film or membrane. Yet another use is as a sealant or component of a sealant to prevent moisture penetration. -23- A solution of the bioadhesive protein can be uni¬ formly coated on a surface as a primer, wuring of the primer coating occurs in a normal air environment by cross-linking, which may be indicated by the develop¬ ment of a brown or tan color when used in high concen¬ tration. A conventional adhesive such as an epoxy adhesive is then applied over the primer coat and the surfaces to be bonded are brought together. In another embodiment of the invention, an adhe¬ sive composition is provided that contains the hydroxylated bioadhesive protein in solution with an¬ other adhesive substance. Typical of the adhesives that may be employed in conjunction with the bioadhe¬ sive protein of the invention are the carbohydrate adhesives and the synthetic resin adhesives such as the polyacrylates, polyepoxides, resols, etc. The known carbohydrate adhesives that can be employed include chitosan, starch, pectin, glucan, dextran, etc. A preferred carbohydrate adhesive is chitosan pur¬ ified from crab or shrimp shell chitin by the proce¬ dure of Skujins, J. J. et aJL. , Arch. Biochem. Biophys. , 111:359 (1965). The free amino groups of chitosan are reactive with the DOPA-derived quinones of oxidized bioadhesive protein, providing covalent cross-links between the two polymers. hitosan at appropriate concentrations provides bioadhesive pro¬ tein mixtures with a high viscosity and excellent adhesive strength. The high viscosity is a particu¬ larly useful property in underwater applications where diffusion can cause a loss of material before the adhesive has an o p portunity to cure. A preferred adhesive mixture comprises from ϊθ.1% to 30% of the hyroxylated bioadhesive polymer and from -24- 1% to 7% chitosan, the balance being solvent. The pH of the composition is from about 5.5 to 7.0. The com¬ position can be cured at pH 6.0 by the addition of catechol oxidase or tyrosinase which catalyzes the formation of DOPA-derived quinones and cross-linking. In another embodiment of the invention, there is provided an adhesive composition in which the bioadhe¬ sive protein is admixed with other proteins that im¬ prove its physical properties such as cohesivity. A preferred protein for this protein is collagen. A preferred composition comprises a solution having 0.1% to 70% solids, the solids in the solution comprising from 1% to 50% bioadhesive protein and from 50% to 99% collagen. The bioadhesive protein of the invention, with or without the addition of other proteins, is parti¬ cularly useful as a biomedical adhesive or sealant in a variety of medical applications, for example, in wound healing. Being a biological material, the bioadhesive protein presents a greatly reduced risk of toxic degradation products -as - compared.._with a synthetic adhesive. The bioadhesive protein can be applied as a biomedical sealant in much the same manner as fibrin (see e.g., Redl, A. and Schlag, C. , Facial Plastic Surgery, 24:315-321 (1985)). The following examples are intended to illustrate further the practice of the invention and are not in- tented to limit its scope in any way. EXAMPLE 1 Production of Antibody to Bioadhesive Precursor Protein Synthetic decapeptide (1.5 mg) with the predomin¬ ant sequence of M. edulis bioadhesive precursor pro¬ tein (ala-lys-pro-ser-tyr-pro-pro-thr-tyr-lys) pre¬ pared by the Merrifield solid state method was combined with 2.0 mg of bovine serum albumin (BSA) in 1.8 ml phosphate-buffered saline. One percent glutar- aldehyde (0.2 ml) was added and the solution was incubated 30 minutes at 22°C. Sodium borohydride was added to a final concentration of 0.5 mg/ml and incu¬ bation was continued at 22°C for one hour. The solu¬ tion was then dialyzed against phosphate-buffered saline. Amino acid analysis of the resulting protein indicated 35 moles of peptide were coupled per mole of BSA. Rabbits were given intramuscular injections with 100 uς of peptide (BSA coupled) incomplete Freund's adjuvant. Booster subcutaneous injections using in¬ complete Freund's adjuvant were given subsequently in two-week intervals. Ant serum with high-titer anti¬ body reactive toward the decapeptide as well as M. edulis bioadhesive precursor protein isolated from mussels or produced in microorganisms was obtained by this method. EXAMPLE 2 Isolation of M. edulis Bioadhesive Precursor Protein mRNA " The adductor muscles of live blue mussels (Mytilus edulis) were cut and the foot organ containing the phenol gland was amputated, quickly cut in small pieces and frozen in liquid nitrogen. The frozen tis¬ sue was shattered into small pieces by mixing in li¬ quid nitrogen at top speed in a commercial Waring blender with a metal container. The pulverized tissue was stored at -80°C until use. The frozen tissue was dissolved in a 4 M guanidine thiocyanate solution as the first step in the RNA iso¬ lation procedure (McCandliss, R. et al., Methods Enzy- 3048 -26- mol, 79_:51-59 (1981)). From 22.1 g wet weight of tis¬ sue, 28.3 mg of total RNA was obtained. Poly A-con¬ taining RNA was purified by selecting the RNA that would bind to oligo-dT cellulose (Aviv, H. and P. Leder, Proe. Natl. Acad. Sci. USA £9:1408-1412 (1972)). Two oligo-dT cellulose selections yielded 0.6% of the total RNA (approximately 170 ug) that in¬ cluded polyadenylated mRNA. As an alternative to the guanidine isolation pro¬ cedure, a modification of the procedure of Berger, S. et al., J. Biol. Chem. , 255:2955-2961 (1980) was used. Five grams of the frozen mussel tissue above after lyophilization were suspended in 100 ml of 10 mM Tris-HCl, pH7.5, 10 mM NaCl, 1.5 mM M Cl 2 , 0.2% NP-40, and 10 mM ribonucleoside-vanadyl complexes (VRC, Bethesda Research Laboratories), .100 ml of phenol were added, and the solution was mixed at top speed in a irtis homogenizer for 2 min. The suspension was centrifuged to separate the phases and. the aqueous phase was removed. The organic phase was extracted with 50 ml of -0.2M sodium .acetate, _pH5.5, and the aqueous phase was combined with the phase earlier recovered. The aqueous phases were extracted three times with phenol containing 0.1% 8-quinolinol. The RNA (with some genomic DNA) was precipitated with ethanol. Poly A-containing RNA was purified as above. From 5 g of tissue, approximately 150 ug of poly A RNA were isolated. -27- EXAMPLE 3 Synthesis and Cloning of M. edulis cDNA Stock Solutions and Materials for First Strand cDNA Synthesis " 0.5 M Tris-BCl, pH 8.3 0.25 M MgCl 2 0.05 M dATP, pH 7.0 0.05 M dGTP, pH 7.0 0.05 M dCTP, pB 7.0 0.05 M dTTP, pH 7.0 32 [alpha- PldCTP, 400 Ci/mmol, 1 mCi/ml (Amer- sha )(Stabilized aqueous solution 0.1 M dithiothreitol (DTT) Oligo ( dT ) 12 _ιo i* 000 ug/ml ( Collaborative Research, W lYham, Mass.) 0.1 M sodium pyrophosphate 0.2 M disodium ethylenediaminetetraacetate (EDTA), pH 8.0 RNasin, ribonuclease inhibitor 30 units/ul, (Promega Biotec) Avian myeloblastosis virus (AMV) reverse tran- scriptase, - approximately . 10,000 units/ml ( obtained- from Life Sciences, Inc., St. Petersburg, Florida) All buffers and salt solutions were autoclaved. The other solutions were prepared with sterile glass- distilled water and were stored in sterile containers. All stock solutions were stored frozen. All enzymes were obtained commercially and used according to the manufacturer's specifications unless otherwise noted. As a template for cDNA synthesis, mRNA prepared by the guanidine method described in Example 2 was employed. In order to follow the synthesis, a radio- -28- 32 active marker ([alpha- PIdCTP) was used. This allows monitoring of all steps by counting Cerenkov radia¬ tion, which does not result in any loss of sample. For each ug of mRNA, 2 uCi of [alpha- 32 ?IdCTP at a specific activity of 400 Ci/mmol were used. The radioactive material was added to a 2X reaction mix¬ ture consisting of 0.1 M Tris-HCl, pB 8.3, 20 M MgCl 2 , 1 mM dATP, 1 mM dCTP, 1 mM dGTP, 1 mM TTP, 20 M DDT, and 5mM sodium pyrophosphate. This solution was kept on ice. To this solution was added mRNA (50 ug/ml, final concentration ) , o igo ( ) ^ 2 « ιg (10 ° ug/ml), RNasin (600 units/ml), AMV reverse transcript- ase (800 units/ml), and enough water to dilute the 2X mix to IX. After 5 minutes on ice, the reaction mix¬ ture was incubated at 46°C for 10 minutes. Following the incubation, EDTA was added to a final concentra¬ tion of 25 mM. The solution was extracted one time with an equal volume of phenol:chloroform (1/1; v/v) and the aqueous phase was chromatographed on a column of Sephadex G-100 (0.7 x 20cm) equilibrated with 10 mM Tris-HCl, pH 8.0, 1 mM EDTA, 0.1 M NaCl. The mRNA:cDNA hybrid in the excluded volume was precipi¬ tated by addition of 0.1 volume of 3 M sodium acetate and 2 volumes of 95% ethanol (-20°C to -80°C). In order to remove the mRNA moiety, the pelleted hybrid was dissolved in 300 ul 0.1 M NaOH and incubated at 70°C for 20 minutes. The solution was cooled on ice and neutralized with 30 ul of IN BC1. The cDNA was precipitated as described above. -29- Stock Solutions and Materials for Second Strand cDNA Synthesis 0.5 M potassium phosphate, pB 7.4 0.25 M Cl j 0.1 M Dithiothreitol (DTT) 0.05 M dATP, pH 7.0 0.05 M dGTP, pB 7.0 0.05 M dCTP, pB 7.0 0.05 M dTTP, pB 7.0 E. coli DNA polymerase I (Klenow fragment), approximately 5,000 units/ml (Boehringer- Mannheim) 10X SI nuclease buffer: 0.5 M sodium acetate, pH 4.5; lOmM ZnSO j , 2 M NaCl, 5% glycerol. It was not necessary to use a radioactive label in the second strand, since the first strand was labeled. A 2X reaction mixture consisting of 0.2 M potassium phosphate, pH 7.4, 20 mM MgCl-, 2 M DTT, 0.4 mM each of dATP, dGTP, dCTP, and dTTP was prepared and kept on ice. To this mixture was added an aqueous solution of cDNA containing the Klenow fragment of E. coli DNA polymerase I (100 units/ml, final concentration), -and. water was added to dilute the reaction mixture to IX. The solution was incubated overnight at 15 C. After the incubation, EDTA was added to 25 mM, the solution- was extracted once with an equal volume of phenol: chloroform (1/1; v/v), and the aqueous phase was chro¬ matographed on a 0.7 x 20 cm column of Sephadex G-100 equilibrated with 10 mM Tris-HCl, pH 8.0, 1 mM EDTA, and 0.1 M NaCl. The DNA in the excluded fractions was precipitated with ethanol as described above. At this point, the ds-cDNA was in the form of a hairpin. The single-stranded loop was removed by di¬ gestion with SI nuclease. The ds-cDNA was dissolved -30- in water and 0.1 volume of 10X SI buffer was added. An appropriate amount of SI nuclease was added and the solution was incubated 20 minutes at 37°C. The amount of enzyme added was determined empirically for each enzyme preparation, since the activity varied from one preparation to another. This was done by measuring the decrease in TCA-precipitable counts from the ds- cDNA. Generally, a decrease of 20-40% was observed. The Sl-digested cDNA was extracted once with phenol: chloroform and the cDNA in the aqueous phase was pre¬ cipitated with ethanol as described above. Addition of Linkers to cDNA For insertion of the cDNA into the EcoRI site of lambda gtlO vector, EcoRI linkers were added to the ends of cDNA molecules. To prevent cleavage of inter¬ nal EcoRI sites in the cDNA molecules with EcoRI, the cDNA was first methylated with EcoRI methylase. Stock Solutions and Materials 5 x EcoRI methylase buffer — 0.5 M Tris-HCl, pH 8.0, 0.05 M EDTA. 8 mM S-adenosyl methionine (SAM) — solution in 0.01 M H 2 S0 4 , pH 2, 10% ethanol. 1 mg/ml bovine serum albumin (BSA) — solution in water, sterile filtered. 8-base pair EcoRI linkers — 10 A 2g0 /ml, obtained from Collaborative Research, Wal ham, Massachusetts. 10 mM ATP, pH 7.0 10 x T4 polynucleotide kinase buffer — 0.7 M Tris-HCl, pH 7.6, 0.1 M MgCl,, 50 mM dithiothreitol. Λ gammat 32Pl-ATP — 10 mCi/ml, 2,000 Ci/mmol, stabilized aqueous solution. 10 x T4 ligase buffer — 0.5 M Tris-HCl, pB 7.8, 0.1 M MgCl 2 , 0.2 M dithiothreitol. 10 x DNA polymerase buffer — 0.5 M Tris-HCl, pB 7.2, 0.1 M MgS0 4 . 2 mM dATP, dCTP, dGTP, dTTP mixture, pB 7.0. 1 M dithiothreitol lOx EcoRI buffer — 1.0 M Tris-HCl, pB 7.5, 0.5 M NaCl, 0.05 M MgCl 2 . The cDNA fragment produced as described above was dissolved in 20 ul of 5 x EcoRI methylase buffer, SAM was added to 80 uM, BSA was added to 0.4 mg/ l, water was added to bring the volume to 99 ul, and 1 ul of EcoRI methylase (20,000 units/ml, New England Biolabs) was added. The reaction was incubated at 37°C for 60 minutes. The reaction was then extracted 2 times with phenol and ethanol-precipitated. The methylated cDNA was collected by centrifugation and dried. Before addition of the linkers, the cDNA was treated with DNA polymerase I (Klenow fragment) in the presence of deoxynucleoside triphosphates to make the ends of the cDNA blunt. The cDNA was dissolved in 24 ul of IX DNA polymerase buffer containing 80 uM each dATP, dCTP, dGTP and dTTP. Two units of DNA polymer¬ ase (Klenow fragment) were added and the reaction mix¬ ture was incubated at 23°C for 10 minutes. EDTA was added to 20 mM and the cDNA was extracted two times with phenol, once with CHC1-, and ethanol-precipi¬ tated. The cDNA, which was EcoRI-methylated and blunt-ended, was collected by centrifugation, washed once with cold 70% ethanol, and dried. S87/03048 -32- To prepare the linkers for addition to the cDNA, the linkers first had to be phosphorylated. 400 Pico- moles of 8-base pair EcoRI linkers were phosphorylated in IX polynucleotide kinase buffer with 20 uCi of [gamma- 32 Pl-ATP and 5 units of polynucleotide kinase. The reaction was incubated at 37°C for 15 minutes. Unlabeled ATP was then added to 1 mM and the reaction was incubated at 37°C for 30 minutes. The enzyme was inactivated by heating the reaction at 65°C for 10 minutes. 160 picomoles of the phosphorylated linkers were then ligated to the cDNA. The blunt-ended, meth¬ ylated cDNA was dissolved in 10 ul of water, 160 pico¬ moles of linkers were added, 10 x T4 ligase buffer was added to IX, ATP was added to 1 mM, and 2 units of T4-DNA ligase (Boehringer-Mannheim) were added. The reaction mixture (a total of 20 ul) was incubated at 15°C for 16 hours. The ligase was inactivated by heating the reaction of 65°C for 10 minutes. At this point, the cDNA had multiple linkers at the ends. Excess linkers were removed by digestion with EcoRI. The ligation reaction was diluted with 5 ul of 10 x EcoRI buffer, 5 ul of 1 mg/ml BSA and 19 ul of water. Prior to addition of EcoRI, 1 ul of the mix was removed for analysis by gel electrophoresis. Ten units of EcoRI (New England Biolabs., 10 units/ul) were added and the reaction was incubated at 37°C for 1 hour. The reaction was extracted 1 time with phenol and run over a column of Sephadex G- 00 equilibrated with 10 mM Tris-HCl, pH 8.0, 1 mM EDTA, and 0.1 M NaCl. The DNA in the excluded fractions was concen¬ trated by use of a Centricon 30 microconcentration -33- unit (Amicon) according to manufacturer's instruc¬ tions. The cDNA was loaded on 5-25% sucrose gradient in 50 mM Tris-HCl, pB 8.0, 1 M EDTA and the gradients were centrifuged at 5°C at 38,000 r.p.m. for 17 hours in a Beckman SW41 rotor. Fractions were collected and were analysed by counting Cerenkov radiation. Paral¬ lel gradients were run with molecular weight markers. Fractions containing cDNA of approximately 1,000-base pairs and longer were pooled and the cDNA was concen¬ trated with a Centricon 30 microconcentration unit. The cDNA was washed four times with 1 ml of H-0 to remove the sucrose and was dried in vacuo. Preparation of Recombinant Bacteriophage DNA Bacteriophage lambda gtlO DNA (Huynh, T.V. et al. , Constructing and screening cDNA libraries in lambda gtlO and lambda gtll. In: D.M. Glover (ed.) DNA cloning. Vol. 1, IRL Press Oxford 1985, pp. 49-78) prepared by standard methods - (Maniatis, et al. , Mole¬ cular Cloning, Cold Spring Harbor, 1982) was linear¬ ized by digestion with EcoRI, phenol-extracted, and precipitated with ethanol. Fifty ng cDNA were mixed with 1 ug EcoRI-cut lambda gtlO (a molar ratio of ap¬ proximately 2:1) in 50 mM Tris-HCl, pH 7, 10 mM MgCl 2 , 20 mM dithiothreitol, and 1 mM ATP, and 1 unit of T4- DNA ligase (Boehringer-Mannheim) was added. The li- gation mixture was incubated for 16 hours at 15°C. A small portion of the reaction mix was analyzed by aga- rose gel electrophoresis. The desired product at this point was high molecular weight, concata eric DNA, which was in the form required for efficient packaging into empty bacteriophage lambda heads. Packaqing of the Recombinant DNA into Bacteriophage Lambda Heads and Transfection into E. coli The recombinant DNA was packaged into bacterio¬ phage lambda heads for introduction into E. coli, by procedures described by Enquist and Sternberg, Methods En∑vmol., _8:281-298 (1979). Packaging extracts are available commercially (Promega Biotec, Madison, Wisconsin, and other sources) and were used according to the manufacturer's instructions. The ligated DNA was mixed with the packaging extracts (50 ul), incu¬ bated at 23°C for 2 hours; the phages were diluted with 0.5 ml, 10 mM MgS0 4 , 10 mM Tris-HCl, pH 7.5, 0.01% gelatin, and a few drops of chloroform were added to the mixture. The packaged phages were stored at 4°C. As a host for titration and propagation of the phage, E. coli strain BNN102 (Huynh, T.V. et al., su¬ pra) was used. On this host strain, only lambda gtlO bacteriophages carrying inserts at the EcoRI site will form plaques. The host strain was grown overnight at 37°C in LB-broth containing 0.2% maltose, to induce synthesis of the phage receptor in the host. The cells were collected by centrifugation and resuspended in one-half volume of 10 mM MgSO.. Cells prepared in this manner were used for 1-2 days. The phage were diluted serially in 10 mM MgSO., 10 mM Tris-HCl, pH 7.5, and 0.01% gelatin. Fifty ul of diluted phage were added to 0.2 ml of E. coli cells, and the mixture was incubated at 37°C for 15 minutes to allow absorption of the phage. The infected cells were mixed with molten (47°C) LB broth containing 0.7% agar and poured on the LB-agar plate. After the top agar hardened, the plates were incubated at 37°C for 5-6 hours, at which time plaques could clearly be seen in the lawn of bacteria. A library of Λ approximately 500,000 recombinant phages was obtained. EXAMPLE 4 Selection of Bioadhesive Precursor Protein cDNA Clones from the M. edulis cDNA Phage Library The lambda gtlO phage library was plated on twenty 15-cm petri dishes at a density of approxi¬ mately 25,000 phage/plate. Duplicate nitrocellulose filter replicates were prepared for hybridization screening (Benton, W.D. and R.W. Davis, Science, 196:180 (1977)). Two 30-base synthetic oligonucleo¬ tides (GCG AAA CCA AGT TAC CCA CCG ACC TAC AAA and GCG AAA CCT TCT TAT CCG CCT ACC TAT AAG) were prepared with an: Applied Biosystems DNA synthesizer. These sequences a e- - two of the possible 98,304 coding sequences for the predominant decapeptide (ala-lys- pro-βer-tyr-pro-pro-thr-tyr-lys) of M. edulis bioad¬ hesive precursor protein. The oligonucleotides were radioactively labeled to a specific activity of approximately 10 8 cpm/ug with gamma-[32P]-ATP (New England Nuclear) and T4 polynucleotide kinase (Boehringer-Mannheim) , then used as hybridization probes. The radioactive oligonucleotides (approxi¬ mately 3.0 ug) were added to 250 ml hybridization solution containing 20% formamide, 6XSSC, 5X Denhardt's solution, 50 mM phosphate buffer (pH 6.8), * 100 ug/ml sonicated denatured salmon sperm DNA and 10% dextran sulfate. The filters were hybridized for 14 hours at 30°C, then washed five times briefly with 300 ml 6XSSC at 22°C, one time with 300 ml IXSSC at 22°C and once at 42°C for 30 minutes with 500 ml IXSSC. The filters were air-dried and autoradiographed at -80°C with Kodak XAR X-ray film for 12 hours. These hybridisation and washing conditions were previously established in hybridisation to M. edulis DNA with the same probe on Southern blots (Southern, E., J. Mol. Biol., 98:503 (1976)>. A 42°C wash in IXSSC was the highest temperature that demonstrated apparently specific hybridization to a fragment of Ba HI-digested M. edulis DNA. The DNA for the South¬ ern blot experiment was isolated from the frozen tis¬ sue described in Example 2 by the method of Blin and Stafford (Blin, N. and D.W. Stafford, Nuc. Acids Res. , 3:2303-2308 (1976)). Plaques giving signals by autoradiography on the duplicate filters were purified by picking, diluting, plating - and repeating the hybridization screen described above. Isolated individual plaques giving radioactive signals were picked and grown as plate lysates for DNA preparation as described (Maniatis, T. et al_. , "Molecular Cloning: A Laboratory Manual," Cold Spring Harbor Laboratory, 1982, pp. 371-372). EXAMPLE 5 DNA Sequence Analysis to Verify Clones DNA from lambda gtlO clone 14-1 and other clones with putative bioadhesive precursor protein cDNA in¬ serts were digested with restriction endonuclease EcoRI. The EcoRI cDNA fragment was cloned into Ml3 mpll (Norrander, J. et al., Gene, 26_:101-106 (1983)). The bacteriophage M13 derivatives were constructed using methods described by Messing (Messing, J., Methods Bnzymol., 101:20-78 (1983)). Both orienta¬ tions of the inserts were represented in independent clones. Some of the larger inserts were progressively shortened by the procedure of Dale et al. {Plasmid, 13:31- 40 (1985)). The phage clones were sequenced by the dideoxy method (Biggin, M.D. et al., Proc. Natl. Acad. Sci. USA, 80:3963-3965 (1983)). The recorded DNA sequences were analyzed with VAX 780 computer assistance. The DNA sequence and translated amino acid sequence of clone 14-1 are presented in Fig. 1. Clone 14-1 encodes a polypeptide with tandem repeats of 19 decapeptides and 1 hexapeptide. Clone 52, presented in Fig. 2, encodes a polypeptide with a mix of 22 decapeptide repeats and 4 hexapeptide repeats preceded by an 82-amino acid segment that is proline rich. Proline-rich segments such as this had not been previously described in M. edulis polyphenolic protein. Clones 14-1 and 52 share considerable homology in their 3*-ends. After a short untranslated trailer sequence in both clones, there is the poly- adenylation signal AATAAA located just prior to a stretch of A residues. These features are expected at the authentic 3.'-terminus of mRNA. EXAMPLE 6 Isolation of Full-Lenqth M. edulis Bioadhesive Precursor Protein cDNA A. mRNA isolation The partial bioadhesive precursor protein cDNAs shown in Figures 1-4 that were isolated and character¬ ized as described in Examples 1-5 are used to isolate -38- full-length bioadhesive precursor protein cDNA. This is accomplished through use of the initial clones as hybridization probes of more carefully prepared cDNA clone banks. The mRNA used in Example 3 was prepared from mussels obtained in a grocery store that were not actively producing byssus threads and adhesive plaques. This may have resulted in low-level and poor quality adhesive protein mRNA. In order to obtain mRNA of better quality, fresh mussels were obtained directly by air freight delivery from the Blue Gold Sea Farms in Rhode Island. The mussels had no treat¬ ment after harvesting other than tumbling to clean their outer shells. The mussels were introduced into a 20-gallon aquarium tank placed in a 4°C room with 10 gallons of synthetic sea water (Instant Ocean, Aquarium Systems, Mentor, Ohio) heated to approximate¬ ly 12-15°C. The tank water was mixed and the temper¬ ature regulated with a Precision H7B heater. Addi¬ tional rapid agitation was provided by a lighting series 30 mixer (Mixing Equipment, Cα.., Inc., Rochester, N.Y. 14603). When individual mussels became active in attachment, they were removed and the distal end of the foot was quickly amputated and immediately frozen by dropping into liquid nitrogen. The tissue was ground into a powder by mixing in liquid nitrogen in a Waring blender, and then lyophi- lized. The mRNA was isolated by the procedure using ribonucleoside-vanadyl complexes as described in Example 2. -39- B. cDNA Synthesis and cDNA Clone Selection The mRNA is used to synthesize cDNA as described in Example 3. However, this time when the double-stranded cDNA is fractionated on a 5-25% sucrose gradient, a much more stringent size selection is used where only cDNA of approximately 3,000 base pairs or larger is pooled for cloning. The phage library and screening is carried out as described in Examples 3 and 4, except that the EcoRI insert fragment of clone 14-1 is isolated and made radioactive by nick translation (Rigby, P.W.J. et ajL., J. Molec. Biol. 1 3:237) with [alρha- 32 P]-dCTP label. Bybridization is carried out at 42°C in 6XSSC, 50% formamide, with 5X Denhardt's solution and 100 ug/ml denatured salmon sperm DNA. The filters are washed at 50°C with 0.1 x SSC with .0.1% SDS. Hybridi¬ zation-positive plaques identified in duplicate . are purified and DNA is prepared as described in Example 4. Clones with the largest insert size are selected for DNA sequence anlysis. Clones found with a charac¬ teristic 5'-terminal sequence and an open reading frame that codes for a protein of approximately 120,000 M.W. are full-length bioadhesive precursor protein cDNA clones. EXAMPLE 7 Construction of Expression Vector PGX2383 for Microbial Production of M. edulis Bioadhesive Precursor Protein The steps outlined below and illustrated sche¬ matically in Figure 6 yielded a plasmid that expresses M. edulis bioadhesive precursor protein as a hybrid fusion protein. The presence of a methionine residue -40- immediately on the amino-terminal side of the adhesive protein allows isolation of basically pure bioadhesive precursor protein sequence after cyanogen bromide cleavage (Gross, 8., Methods Enzvmol. r 11s238 (1967)). Plasmid pGX2287 CSRRL-B15788), part of a highly optimised vector/host system for expression of bovine chymosin (see copending U.S. Patent Application Nos. 671,967 and 644,998) was used as the starting vector. The EcoRI fragment from lambda gtlO clone 14-1 was subcloned into the EcoRI site of plasmid pGX2627. Plasmid pGX2627 is derived from pGX1066 (on deposit at the American Type Culture Collection, Rockville, Mary¬ land, and having accession number ATCC 39955). To derive pGX2627, a 514-base pair Rsal fragment from wild-type phage M13 containing the M13 origin of rep¬ lication was inserted into the EcoRV site of pGX1066. The DNA sequence of pGX2627 is shown in Figure 7. ) This plasmid contains a multi-restriction site sequence as shown in Figure 5(a). Ten icrograms of plasmid pGX2627 DNA and 20 micrograms of lambda clone 14-1 DNA were cut with EcoRI (New England Biolabs). This enzyme reaction and all those outlined below were performed according to the manufacturer's recommenda¬ tions. The pGX2627 DNA was then treated with alkaline phomphatase. After phenol-chloroform extraction and ethanol precipitation using standard procedures in the art, a ligation using T4 DNA ligase in 20 ul with 1 ug of the EcoRI-cut, phosphatase-treated pGX2627 DNA and 12.5 ug EcoRI-treated lambda 14-1 DNA was started at 15 C. After 30 minutes, the reaction mixture was di¬ luted to 50 ul and ligation was continued for nine hours. The ligation mix was used to transform E. coli 41 DHl (D. Banahan, J. Mod. Biol. 166:557-580, 1983) us¬ ing standard procedures (Mandel, M. and Biga, A., J. Mol. Biol. , 53:154, 1970). Most of the transformants contained the desired insert. One recombinant plasmid was identified as pGX2368 as shown in Figure 6. Plasmid pGX2346 was constructed from pGX2287 by insertion of a synthetic oligonucleotide (Fig. 5b) between the Clal and SphI sites of pGX2287. Plasmid PGX2287 DNA was cut with Clal and ligated with synthetic DNA at high molar concentration. The ligated DNAs were then cut with SphI and ligated again at low concentration to form circles. Plasmid pGX2346 was identified by its decreased size and verified by DNA sequencing. An in-frame gene fusion between the trpB portion of pGX2346, and the bioadhesive precursor protein cDNA of pGX2368 is constructed in the following manner: A Bell endonuclease recognition site was first placed at the translation stop.codon of pGX2368 by changing two bases as -indicated in Figure.1 using oligonucleotide- . directed mutagenesis (Zoller, M.J. and M. Smith, Meth- . ods Enzymol., .100:457-500, 1983) to create plasmid pGX2380. Both plasmids pGX2380 and pGX2346 are grown, for DNA preparation in an E. coli host that contains the dam mutation (defective in DNA adenine methylase) so that they could be digested with Bell. The non-methylated pGX2346 DNA is cut with NotI and the pGX2380 DNA is cut with Xbal. Then both DNAs are treated with E. coli DNA polymerase (Klenow fragment) to fill in the 5' single-stranded DNA overhangs. The DNAs are then ligated at high DNA concentration (approximately 2 ug of each DNA in 20 ul) with T4 li- gase. The ligation product is cut with Bell then ligated again at low DNA concentration (approximately 1 ug total DNA in 150 ul volume) and used to transform E^ coli GX3015. A transformant with the desired con¬ struction (see Figure 6) is designated pGX2383. GX3015 cells with plasmid pGX2383 produce a bioad¬ hesive precursor protein of approximately 24,000 M.W. upon induction of the hybrid lambda promoter by a shift of growth temperature from 32 to 37°C. EXAMPLE 8 Construction of Expression Vector pGX2381, for Microbial Production of M. edulis Bioadhesive Precursor Protein The EcoRI insert fragment from lambda gtll clone 52 (see Fig. 2) was cloned into the EcoRI site of plasmid pGX2627. Plasmid pGX2627 is derived from pGX1066 that has been deposited at the American Type Culture Collection (ATCC. 39955). To derive plasmid pGX2627, a 514-base pair Rsal fragment from wild-type phage M13.containing the M13 origin of replication was inserted into the EcoRV site of pGX1066. The DNA se¬ quence of pGX2627 is shown in Figure 7. Ten m crogr*ms of plasmid pGX2627 DNA and twenty micrograms of lambda clone 52 DNA were cut with EcoRI. The pGX2627 DNA was then treated with alkaline phos- phatase. After phenol-chloroform extraction and etha¬ nol precipitation using standard procedures in the art, one microgram of phosphatase-treated pGX2627 DNA and twelve micrograms of the EcoRI-cut lambda clone 52 DNA were ligated at 15°C in a twenty microliter volume with T4 polynucl<»otide ligase. After thirty minutes the ligation mixture was diluted to fifty microliters with ligation buffer and the reaction was continued for nine hours. The ligation mix was used to trans¬ form E. coli DH1 (D. Hanahan, J. Mol. Biol. 166:557-580, (1983)) using standard procedures (M. Mandel and A. Biga, J. Mol. Biol. 53:154, (1970)). Most of the transformants contained the desired insert. One recombinant plasmid with the cDNA inserted in the orientation depicted in Figure 6 was identified as pGX2377. A Bell endonuclease site was placed at the translation stop codon of the cDNA in pGX2377 by changing the two bases as indicated in Figure 2 using oligonucleotide-directed mutagenesis (J.M. Zoller and M. Smith, Methods Enzymol. 100:457-500, (1983)) to create pGX2381. The mutagenic oligonucleotide. had. the. DNA sequence ACAGATATTGATCACAATATTA. EXAMPLE 9 Yeast Expression of Bioadhesive Precursor Protein (Clone 14-1) YpGX265GAL4 is on deposit at the American Type Culture Collection, Rockville, Maryland, having ac¬ cession number ATCC 67233. The YpGX265GAL4 vector contains the following functional units: Promoter - Hybrid derived from the S. cerevis¬ iae GAL1 and MF-alpha 1 (alpha-factor) promoters. Permits galactose-regulated expression. Regulatory gene - GAL - encodes GAL4 protein, a positive regulator of the GAL -MF-alpha 1 hybrid pro¬ moter. Derived from S. cerevisiae. Terminator - derived from synthetic DNA-based on S. cerevisiae GAPDH transcription terminator. Signal encoding sequence - Derived from synthetic DNA- based on S. cerevisiae PB05 signal. Codons designed substantially for usage preference in S. cerevisiae. LEU2-d gene - Marker for plasmid selection in S. cerevisiae. Derived from plasmid pJDB207. 2-micron DNA - Derived from plasmid pJDB207. Provides plasmid replication origin for S. cerevisiae. _!• coli replication origin - Derived from pJDB207. E. coli selectable marker - Ampicillin resis¬ tance - derived from pJDB207. Foreign genes are inserted between PH05 signal encoding sequence and the GAPDH terminator. 1. YpGX265GAL4 is digested with restriction endonuclease Hindlll and the large and small DNA frag¬ ments are gel-purified. The small (3.65 Kb) fragment is saved for a later, step.. 2. The large Hindlll fragment is digested with restriction endonuclease Smal, generating two DNA fragments. 3. M13mp9 (commercially available) is digested with Smal and Hindlll and then treated with calf alka¬ line phosphatase. 4. The DNA molecules generated in steps 2 and 3 are ligated and used for transformation of E. coli. The transformants containing phage double-stranded DNA with the yeast promoter-terminator cassette are iden¬ tified. 5. The recombinant phage double-stranded DNA is digested with restriction endonucleases EcoRV and BamHI and the large fragment (approximately 8kb) is gel-purified. 6. Plasmid pGX2380 (prepared in accordance with Example 7 and containing bioadhesive precursor protein cDNA) is digested with restriction endonuclease Xbal, and the single-stranded overhang is filled with the Klenow fragment of DNA polymerase. 7. The DNA fragment generated in step 5 is di¬ gested with Bell. The small DNA fragment (approxi¬ mately 625bp) is gel-purified. 8. Two oligonucleotides of sequence: 5' ATCAAATCGATGGCGGCC and 5* GGCCGCCATCGATTTGAT are annealed and used in a three-way ligation with the DNA fragments generated in steps 5 and 7. 9. E. coli transformants containing- the desired phage DNA molecules are identified by restriction en¬ donuclease digestions ■ ■ - - - 10. The phage DNA generated in step 9 is digested with restriction endonucleases Smal and Hindlll and the small fragment (approximately 1300bp) is gel-purified. 11. YpGX265GAL4 is digested with Smal and Hindlll and the large fragment is gel-purified. 12. The DNA molecules generated in steps 10 and 11 are ligated and E. coli is transformed. The de¬ sired plasmid, YpGX285, is identified by restriction endonuclease digestion. 13. YpGX285 is linearized with restriction en¬ donuclease Hindlll and treated with calf alkaline phosphatase. 14. The 3.65 kb GAL4-containing fragment from step 1 is ligated with the linearized YpGX285 generated in step 13. 15. E. coli is transformed and the desired plasmid, YpGX285GAL4, is identified by restriction endonuclease analysis. 16. Saccharo yces strains carrying mutations in the LEU2 structural gene are transformed with YpGX285GAL4 by standard methods. A suitable strain is AH22 (ATCC #38626). 17. The yeast strain is grown in YNBD medium to maintain the plasmid. The YNBD medium contains 0.7% yeast nitrogen base, 2% glucose, and 20 mg/liter L-histidine. 18. For production of the bioadhesive precursor protein the strain is grown in medium containing 1% yeast extract, 2% peptone, 1% glucose, and 1% galac- tose at 30°C with shaking. 19. Production of the bioadhesive precursor protein is analyzed by Western Blot and SDS-polyacryl- amide gel electrophoresis of the yeast proteins following procedures known in the art. EXAMPLE 10 Growth of GX3015 (PGX2383) and Induction of Bioadhesive Precursor Protein Synthesis Cultures are maintained using growth tempera¬ tures of 30°C with ampicillin at 100 ug/ml and/or med¬ ium lacking tryptophan to maintain nutritional selec- -47- tion. Plasmid pGX2383 contains the bla, gene which en¬ codes beta-lactamase providing ampicillin resistance, as well as the trpED that in tryptophan-deficient med¬ ium complements the trpED102 deletion in the GX3015 chromosome. A single colony of GX3015 (ρGX2383) is picked after growth on minimal salts medium (Miller, J.B., "Experiments in Molecular Genetics," Cold Spring Barbor Laboratory, 1972, p. 432) supplemented with 0.4% casamino acids and 0.4% glucose and inoculated into 5 ml of LB medium supplemented with 100 ug/ml ampicillin. After reaching an optical density Ag 0Q of greater than 1.0, 0.4 ml of the culture is inocu¬ lated into each of two 250-ml baffled flasks contain¬ ing 50 ml of LB broth supplemented with 100 ug/ml ampicillin. The two flasks are incubated at 30 C and shaken at 250 r.p.m. for 6.5 to 9 hours. Fermentation is carried out using 8 liters of the following initial medium: (NH 4 ) 2 S0 4 - 30 g KH 2 P0 4 - 15 g K 2 HP0 4 - 5 g Biotin (0.5 mg/ml in 95% ethanol) - 12 ml tap water to 8 liters, autoclave The following additions are made after autoclaving to provide the initial medium: CaCl 2 .2H 2 0 - 10 ml of 10% (w/v) sterile solution glucose - 360 ml of 50% (w/v) sterile solution niacin - 18 ml of 0.5% (w/v) sterile solution Trace solution 1 - 90 ml Trace solution 2 - 18 ml Trace solution 3 - 1.8 ml The following fermentation conditions are main¬ tained: pH 7.0 (controlled by 5N NH 3H, and IN H 3 P0 4 ) Sparge rate - 1 vvm Temperature - 32 C Agitation rate - 800 r.p.m. In order to increase cell density prior to induc¬ tion of expression, a system of broth supplementation with nutrients is undertaken. The feed solution is prepared as follows: 1000 g glucose with deionized water for final vol¬ ume of 1700 mis is autoclaved. After autoclav- ing, there is added: Trace solution 1 _ 1 500 ml Trace solution 2 LOO ml Trace solution 3 - 10 ml CaCl 2 .2H 2 0 — 50 ml Trace Solution tl FeCl,.6H,0 - 5.4 g ZnSOT.7H,0 - 1.44 g MnCl*.4H,0 - 1.0 g CuSO^.5H,0 - 0.25 g CoCl,.6H,0 - 0.24 g H 3 B0 3 - 0.062 g Brought to 1000 ml and sterile filtered, Trace Solution t2 H-0 - 900 ml HCl - 44.8 ml MgS0 4 .7H 2 0 - 61.6 g Brought to 1000 ml and sterile filtered. -frace Solution 43 B,0 - 1000 ml Na 2 Mo0 4 .2B 2 0 - 24.1 g Sterile filtered. The feed solution is initially added to the broth in a volume of 180 ml and thereafter as needed to maintain the glucose level at 10 g/liter. Feed sup¬ plementation is continued until the A g00 reaches 20, at which time the cells are induced to express bioad¬ hesive precursor protein from the hybrid lambda promoter. Induction is effected by raising the tem¬ perature to 37°c to deactivate the temperature-sen¬ sitive lambda cI857 represser protein produced by the defective lysogen in the GX3015 chromosome. The fermentation is maintained at 37°C for another 6-8 hours. EXAMPLE 11 Purification of Bioadhesive Precursor Protein E. coli GX3015 (pGX2383) cells (32 g wet weight) are suspended in 20 ml 20 mM Tris-HCl, 2mM EDTA (pH 7.5), 1 mM phenylmethylsufonyl fluoride, 25 mM iodo- acetic acid, and thoroughly disrupted by passage through a French press followed by sonication. The cell debris and protein-containing inclusion bodies containing bioadhesive precursor protein are pelleted by centrifugation at 27,500 g for 30 minutes at 4°C. The pellet is extensively washed by suspension in 10 mM Tris-HCl, 1 mM EDTA (pH 7.5) and centrifugation. Washing is continued until the supernatant is clear. The pellet is then dissolved in 15 ml of 6 M guanidine hydrochlor de, 5% beta-mercapto thanol t 25 M iodo- acetic acid, and centrif ged at 30,000 xg for 30 minutes at 4°C. The supernatant is dialyzed against 4 liters of 0.2 mM EDTA, 10 mM iodoacetic acid, with 3 changes which results in protein precipitation. The precipitate containing about 0.5 g of protein is dissolved in 40 ml of 70% formic acid. Cyanogen bromide (1 * 3 g). is added and the solution is allowed to react overnight at room temperature. After rotary evaporation, the residue is extracted with 20 ml water (pH 4.0 from residual formic acid). The pH of the water-soluble fraction is adjusted to pH 7.0 with 5N KOH resulting in some precipitate formation. The supernatant is then applied to a CM cellulose or S-Sepharose column (1.5 x 26 cm) equilibrated with 50 mM potassium phosphate (pH 7.0). After the column is washed for 14 hours with 50 mM potassium phosphate, the bioadhesive precursor protein is eluted with either a salt gradient (0 to 0.5 M KCl) or a pH change (pH 8 to pH 10) in the buffer. The fractions are assayed by measurement of absorbance at 280 nm and by SDS polyacrylamide gel electrophoresis using both coomassie blue protein stain and the Western blot assay with specific -antibodies (Example L) . The fractions containing the bioadhesive precursor protein are pooled and dialyzed overnight twice against 2 liters of deionized water. The resultant suspension is lyophilized and 1 mg of purified material is obtained. Material could be further purified, if necessary, using Sephadex G-75 column chrom tography with Q3M ammonium acetate pH 4.0. Folyphenolic protein eluting in the first protein peak is dialyzed against water and lyophilized for recovering as a salt-free powder. The purified protein is hydrolyzed in 6 M constant boiling BC1 with phenol crystal i i vacuo at 105°C for 24 hours. The amino acids in the acid hydrolysate are identified as O-phthaldehyde (OPA) derivatives which are separated on C. g reverse-phase BPLC column (Fleury, M.O. and D.V. Ashley, Anal. Biochem. , 133:330-335 (1983)). The amino acid composition is used to verify purity since only a subset of amino acids is present in bioadhesive precursor protein. EXAMPLE 12 Alternate Method for Bioadhesive Precursor Protein Purification E. coli cells [GX3015 (pGX2383)l or yeast cells tAH22(YρGX285GAL4H from a 180 1 fermentation are centrifuged in a Westphalia centrifuge, washed with saline and resuspended as a 30% solids suspension in 10 mM EDTA, 1.0 mM phenylmethylsul onyl fluoride (PMSF), 10 mM iodoacetic acid (IAA) at pH 8.0 before breaking by a Manton-Gaulin homogenizer. Bioadhesive precursor protein present in an insoluble fraction is collected with the cell debris upon Westphalia centri¬ fugation. The centrifugation pellet is made . a 20% solids suspension in acetic or formic acid at pH 2.2 to 2.5 and mixed for several hours to solubilize bioadhesive precursor protein. The extract is centri¬ fuged or filtered to remove solids and the clear filtrate solution is then adjusted to pH 4 with 5 N KOH in the presence of 10 mM EDTA, 10 mM IAA and 1.0 mM PMSF. Impurities are removed as a precipitate by filtration or centrifugation. The polyphenolic protein in the clear supernatant is then concentrated by ultrafiltration (10,000 M.W. cutoff membrane) and subsequently lyophilized. After the residue (approxi¬ mately 50-100 g) is dissolved in 1.2 liter of 70% formic acid, cyanogen bromide (100 ς) is added and the solution is stirred for 24 hours at room temperature. The reaction mixture is then dried by rotary evapora¬ tion and the residue is dissolved in 50-100 ml 6 M guanidine hydrochloride at pH 8.0 and centrifuged at 30,000 g for 20 minutes at 4°C. The supernatant is chromatographed on a Sephacryl S-300 column equili¬ brated with the same solution. The fractions contain¬ ing bioadhesive precursor protein are collected, adjusted to pH 4.0 with acetic acid, and dialyzed against water and recovered as a salt-free powder. EXAMPLE 13 Hydroxylation of Bioadhesive Precursor Protein Since tyrosinase has been known to catalyze the hydroxylation of tyrosine and oxidation of DOPA (Ito et al., Biochem., 2_2_2:407-411 (1984); Maru o and Waite, Biochem. Biophys. Acta, 8_92:98-103 (1986)), mushroom tyrosinase or Streptomyees antibiσticus tyrosinase can be used to enzymatically modify the homogenous E. coli or yeast-produced bioadhesive precursor protein. To a 1 ml mixture containing a 2 mg protein, 25 umole ascorbic acid and 0.05 M sodium phosphate between pH 5 to 7.5, 0.1 mg mushroom tyrosinase (Sigma Chemical Co.) is added. The mixture is allowed to react at room temperature for 3 hours. The kinetics of the hydroxylation process can be monitored by the colorimetric assay for DOPA and DOPA-derived quinone (Waite, J.H. and M.L. Tanzer, Anal. Biochem., 111:131-136 (1981)). The product is further analyzed by amino acid analysis after acid hydrolysis as above (Example 9). After correcting for loss during the recovery process, amino acid analysis indicates approximately 40% of the tyrosine residues are converted to DOPA. After hydroxylation, the pH of the solution is adjusted to 4 with acetic acid and the solution is dialyzed against 100 volumes of 5% acetic acid. The samples are rotatory-evaporated to reduce the volume. The tyrosinase is removed either by using a LH-Sepha- dex 60 column, which is eluted with 0.2 M acetic acid, or using a membrane filtration method (Amicon PM30, cut off of 30,000 M.W. ). An alternative purification scheme after hydroxy¬ lation is described below. After CNBr cleavage, the supernatant obtained at pH 7.0 (see Example 11) is acidified to between pH 5 and 7 in the presence of ascorbic acid or tropolone (Kahn, V. and A. Andrawis, Phytochemistry, 24:905-908 (1985)). Hydroxylation is started by the addition of tyrosinase. At the end of the reaction, the sample is purified by an SE Sephadex column. The fractions containing DOPA are pooled, dialyzed against 2.5% acetic acid and lyophilized. The purity of the hydroxylated protein is established by acid-urea polyacrylamide gel electrophoresis (Panyium S. and R. Chalkley, Arch. Biochem. Biophys. , 130:337-346 (1969)) and amino acid analysis. The DOPA-containing bioadhesive protein is then ready for formulation as an adhesive. -54- EXAMPLE 14 Use of Bioadhesive Protein as a Primer for Conventional Adhesives Surfaces such as metals or plastics are frequently given a pretreatment such as oxidation with acid, flame treatment or plasma bombardment to improve the ability of a surface to "wet"* or interact with the adhesive. Microbially produced and hydroxylated mussel bio¬ adhesive protein coated onto a surface can be used as a pretreatment or priming substance for conventional adhesives. An example of the use of mussel bioad¬ hesive protein as a primer treatment for bonding two pieces of aluminum is given below. Hydroxylated bioadhesive protein prepared as in Example 13 is dissolved in degassed water (optimally at pH 7.0 to 8.0) at a concentration of 10-400 mg/ml (10-40% w/v). The solution is maintained under nitro¬ gen to prevent premature oxidation of DOPA residues to quinones and curing of the adhesive primer. The bioadhesive protein solution is sprayed or painted to uniformly moisten -an oil-free aluminum sur¬ face in a normal air environment. The surface is then dried in a low-humidity environment. A brown or tan color may develop indicating quinone oxidation and chemical cross-linking. The primed surfaces to be bonded are then joined using standard materials such as epoxy glue. As an alternative to speed bioadhesive protein curing and eliminate the prehydroxylation step (see Example 13 ) , an enzyme such as mushroom tyrosinase ( Ito et al.. Biochemistry, 222:407-411 (1984)) or Streptomvces tyrosinase (Lerch and Ettlinger, Eur. J. Biochem., 31:427-437 (1972)) is mixed with the non-hy- droxylated protein (Example 11) immediately prior to application (for example, in the nozzle of a spray applicator) at a concentration of 0.01 to 1.0 mg/ml solution. The enzyme under these conditions effects oxidation all the way from tyrosine to the reactive quinone species. Blends of bioadhesive protein with other polymers (as outlined in adhesive examples given below) are also used as primers for other adhesives. EXAMPLE 15 Use of Bioadhesive Protein The hydroxylated bioadhesive protein prepared as in Example 13 is dissolved at a concentration of 1- 700 mg/ml (0.1-70% solids) in water (or a physio¬ logical salt solution for medical applications) ad¬ justed to pH 6.0 with dilute acids. Immediately prior to application to a surface, a basic solution (approximately 1/50 volume) is added to increase the pH to 8.0. An enzyme such as mussel catechol oxidase (Waite, J.H., J. Mar. Biol. Assoc. , 6_5:359-371 (1985)) can also be added (final concentra¬ tion 0.01-1 mg/ml) in place of or in addition to base solution immediately prior to application to acceler¬ ate the oxidation of DOPA residues to quinones to yield more rapid curing. Mixing of components immedi¬ ately prior to application can occur, for example, in a spray head of a Duploject syringe as has been de¬ scribed for fibrin sealant (Redl, H. and G. Schlag, Facial Plastic Surgery, 2:315-321 (1985)). Alterna- tive enzymes such as mushroom tyrosinase (Ito et al^ , Biochemistry, 222:407-411 (1984)) or Streptomyces ty¬ rosinase (Learch and Bttlinger, Eur. J. Biochem., 33.: 27-437 (1972)) are also used. With tyrosinase, the bioadhesive protein need not be previously hydroxylated and thus the material without prior hydroxylation described in Example 11 may be used. In order for oxidation of DOPA residues to quinones to take place and subsequent curing (either with or without enzymes), there must be dissolved oxygen present in the bioadhesive protein solution. EXAMPLE 16 Composites of Bioadhesive Protein with Other Protein Polymers In order to moderate and improve the properties of the mussel- polyphenolic protein adhesive, blends with other polymers are used. The bioadhesive protein is naturally associated with collagen in the byssal threads of the mussels. Collagen is one natural polymer that can be used to increase the cohesive strength of phenolic protein composites. Acid-soluble collagen (Gallop, P.M. and S. Seifter, Methods Enzymol. , VI:635-641 (1963)) is dissolved in dilute acid solution at 10-70% (w/v) . The collagen is mixed with the bioadhesive protein mixtures as described in Example 15 in ratios having from 1% to 50% of the solids comprising bioadhesive protein with total solids ranging from 10 to 70%. Higher percentages of bioadhesive protein yield more highly cross-linked rigid composites than those with lower percentages of bioadhesive protein. Alkaline solution may be used to neutralize the mixture immediately prior to applica¬ tion. This allows more rapid oxidation and cross-linking (curing) of the mixture. Also, at neutral pH, the collagen will crystallize providing added cohesive strength. Alternatively, the bioadhesive protein is used in combination with preformed sheets of collagen. This method is analogous to the use of reinforcing steel in cement or graphite fiber in epoxy composites. Collagen sheets such as the commercially available collastat (a Helitrex product distributed by American Home Products Corporation) or other similar products are sprayed or soaked in bioadhesive protein activated as described above in Example 13, then applied to the surface to be bonded. In a similar manner, other types of insoluble or crystalline protein sheets can- be used as reinforce¬ ment for adhesive protein. For example, silk cloth, or sheets formed from solubilized and reprecipitated alpha-keratose from wool keratin fibers (J. De Bersag- bes, Curr. Probl. Dermatol. , 6_:34-86 (1976)), or poly¬ merized fibrin clot formed from purified fibrinogen, thrombin and Factor VIII (Redl, H. and G. Schlag, Facial Plastic Surgery, -2 :315 *" 321 (1985)) are used. For medical applications, the use of sheets of fibrin may have the additional benefit of helping to promote wound healing (Redl and Schlag, supra). EXAMPLE 17 Composites of Bioadhesive Protein with Carbohydrate Polymers Chitosan is dissolved in 1% acetic acid to a con¬ centration of 30-150 mg/ml (3-15% w/v). The chitosan solution with a final pH of approximately 6.0 is blended with the bioadhesive protein solution described in Example 15. Blends typically have bioad¬ hesive protein concentrations between 2 and 30% and chitosan concentrations between 1 and 7%. At pH 6.0 where the chitosan is still soluble, catechol oxidase and tyrosinase addition catalyzes the formation of reactive DOPA-derived quinones and cross-linking. Increasing the pH to 8.0 as in Example 15 prior to application results in the immediate precipitation of chitosan out of solution which in some cases may not be desirable. EXAMPLE 18 Isolation of DNft Clone Nl Encoding M. edulis Bioadhesive Precursor Protein Messenger RNA was isolated from fresh _V_ edulis as described in Example 6. Two 18-base probes were synthesized based on the DNA sequence in clone 14. The first synthetic probe (#2214 - GTTTGTTGGTTTATATGC) was complementary to a conserved sequence close to the 3'-end of the coding sequence. This oligonucleotide was also used to prime cD A synthesis. The second probe (#2213-TTTATAAGTTGGCTTTGC) was the complement of the sequence that codes for the hexapeptide found in several locations in the cDNA clones. Probes 2213 and 2214 were used together since they have the same GC/AT ratio and thus approximately the same melting tempera¬ ture. The use of both probes provided multiple hybridization points and a strong signal during clone bank screening. The clone bank prepared as in Example 6 was screened with the 18-base oligonucleotides. Thirty hybridization-positive clones were initially identi¬ fied in duplicate. Twenty-eight of those were plaque purified with two rounds of plating and screening, and their DNA was analyzed. A third synthetic 18-base probe (#2231-GGGATATATTGACTTGGA) , which encodes a portion of an uncommon, variant decapeptide, was included in this hybridization analysis. As expected, many of the clones had larger cDNA inserts than in the cDNA cloning experiments described in Example 4. The largest clones, for example N5, N14 and N26, had a primary insert fragment of approxi¬ mately 2.6 kb. Clones Nl, N14 and N15 were chosen for initial DNA sequence analysis because these DNA pre¬ parations were the cleanest. Clone Nl had the largest insert that hybridized to all three probes and it displayed an intense signal with probe #2213, which encodes the predominant hexapeptide sequence. An initial sequence analysis of two of the largest clones <N15 and N14) showed that both were false posi¬ tives for polyphenolic protein sequence. The 2.1 kb EcoRI insert from clone Nl was sequenced as described in Example 6, with the following modi ications. The Nl clone could be subcloned stably in only one orien¬ tation in the M13 sequencing vectors and the progres¬ sive sequencing strategy of Dale et al. ,Plasmid, Vol. 13, p. 31-40, (1985) was performed on only one strand. To confirm that the sequence was assembled correctly, the inserts from the series of overlapping subclones from the Dale procedure were sized by electrophoresis on agarose gels. DNA sequencing confirmed that clone Nl contains polyphenolic protein cDNA. In fact, this clone appears to encode a very large middle portion of the mussel adhesive protein and contains 76 continuous tandem repeats of decapeptides and hexapeptides throughout the 2.1 kb length of the clone (Figure 9). Gaps in the sequence shown in Figure 9 allow optimal alignment of the hexapeptides with the decapeptides. Of the 63 decapeptides in the Nl protein, 31 are identical to the decapeptide described in U.S. Patent 4,585,585. The variations in each position are shown in Figure 10. Of the 13 hexapeptides, 10 are Ala-Lys-Pro-Thr-Tyr-Lys, two have Ser instead of Ala, and one has Val instead of Ala. Clone Nl does not have a perfect overlap with any of the other cDNA clones sequenced to date. While it is possible that Nl lies just upstream of carboxy-ter- minal clones 52 or 14, it is also possible that the lack of perfect overlap in any of these clones is the result of multiple polyphenolic protein genes or alternate splicing patterns, as found in the studies of other structural proteins. Clone Nl also does not have a characteristic 5'-end sequence and therefore probably does not encode the N-terminus. However, it encodes a polyphenolic protein of nearly 80,000 molecular weight and therefore constitutes the major¬ ity of the 110,000-130,000 intact mussel protein. Except for the proline-rich segment of clone 52, these sequencing results suggest that the tandem deca- and hexapeptide repeats may constitute the complete functional part of this protein with respect to adhe¬ sive properties. One could easily increase the size of the encoded protein by ligating two or more of the sequenced cDNA segments together. EXAMPLE 19 Expression of Nl cDNA in Yeast For expression of the Nl cDNA in yeast, it was desirable to position NotI and BamHI restriction endo¬ nuclease sites at the 5* and 3' ends of the coding sequence respectively. To accomplish this, vector M13mpl8 was first mutagenized by oligonucleotide directed mutagenesis to insert a NotI site adjacent to the unique EcoRI site. An oligonucleotide of sequence. EcoRI NotI 5 ' CCGAGCTCGAATTCTCGCGGCCGCGTAATCATGGTCAT was used to prime this mutagenesis. The new M13 vector was called MGX463 and has unique NotI and BAMHI restriction sites bordering the EcoRI site. MGX463 double-stranded DNA was digested with EcoRI and treated with calf-alkaline phosphatase and the linear DNA was purified on a 0.8% agarose gel. The Nl cDNA coding sequence was removed from an Ml3mpl0 vector by digesting double-stranded DNA with EcoRI. The approximately 2200 base pair EcoRI frag¬ ment was gel purified. This fragment was ligated with the MGX463 double-stranded DNA which was linearized with EcoRI and treated with calf alkaline phosphatase. E. coli strain GX1210 was transformed with the liga¬ tion mix. This resulted in vector MGX464 which carries the Nl cDNA sequence bordered by NotI and BamHI restriction sites. MGX464 was digested with NotI and BamHI and the Nl cDNA fragment was gel purified. YpGX283 (fully described in commonly assigned co-pending U.S. Patent Application Serial No. 025,243, filed March 12, 1987, and shown in Figure 11 ) was digested with NotI and BamHI and the large vector fragment was gel purified. This fragment was ligated with the Notl/BamHI Nl cDNA fragment and E. coli strain GX1210 was transformed, resulting in plasmid YpGX405. Plasmid YpGX405 (Figure 12) is a yeast E. coli shuttle vector which contains the Nl cDNA sequence positioned between the PH05 signal coding sequence and the GAPDH transcription terminator. Expression is directed by a hybrid promoter composed of elements of the yeast GAL1 and MF-alpha 1 pro¬ moters. In order to insert the GA gene into YpGX405, that plasmid was linearized with Hindlll and treated with calf alkaline phosphatase. Vector YpGX283GAL4 (fully described in co-pending commonly assigned U.S. Patent Application Serial No. 025,243, filed March 12, 1987), was digested with Hindlll and the GAL4 gene was gel purified. These two fragments were ligated and E. coli strain GX1210 was transform¬ ed, resulting in YpGX405GAL4. To determine if S_. cerevisiae produced bioadhesive precursor protein of the expected molecular weight, yeast strain D8 transformed with expression vector YρGX405GAL4 first grown on YNBD solid medium (0.7% yeast nitrogen base, 10% glucose, 2% agar) and then inoculated into 10 ml YPD (1% yeast extract, 2% peptone, 2% glucose) so that the initial A fi0 _ reading was 0.1 and was then grown at 28°C with shaking for 17-24 hours. The cells were harvested and washed with 10 ml YPGal (1% yeast extract, 2% peptone, 2% galac- tose) and resuspended in an equal volume of YPGal and induced for 6-28 hours. One ml of that culture was harvested and washed with T 25 E 125 pH 8.4 buffer (25 mM Tris-HCl, 125 mM EDTA, pH 8.4). The cells were then resuspended in 100 ul T 25 E 125 buffer, and broken by vortexing in the presence of glass beads. Follow¬ ing the addition of 200 ul T 2 _ E. 25 , the cell lysate was removed from the glass beads and cell debris was pelleted in a microfuge for five minutes. The insol¬ uble pellet was resuspended in 200 ul sample buffer (Laem li, U.K. 1970, Nature 227' 68 °- 685) and boiled for five minutes. A 25 ul aliquot was examined on a 10% SDS-polyacrylamide gel and stained with Coomassie blue. The results of this analysis showed that bioad¬ hesive precursor protein of the appropriate molecular weight was produced by the yeast strain at levels of approximately one percent of the total cell protein. EXAMPLE 20 Purification of Bioadhesive Precursor Protein Yeast cells as a 30% suspension in 20 mM Tris pH 7.5, 2 mM EDTA, 0.1 mM PMSF, 10 mM iodoacetic acid were thoroughly disrupted mechanically by a French Press, sonicator or a Manton Gaulin homogenizer. The adhesive precursor protein present in the insoluble fraction was collected with cell debris by centrifuga¬ tion at 25,000 g for 30* at 4°C. The pellet was extensively washed with 10 mM Tris, 1 mM EDTA (pH 7.8) and centrifuged as before. The pellet was resuspended as a 30% suspension in formic acid (final concentra¬ tion of 70%) and mixed for several hours. Cyanogen bromide (50 gm/1) was added and the suspension was- stirred for 24 hours at room temperature. The reac¬ tion mixture was then rotary evaporated to dryness, and the residue was resuspended to the original lysis volume with water (pH 3.0) and stirred for several hours to extract the adhesive protein. After centrifugation (as before) , the adhesive precursor protein was precipitated from the superna¬ tant with 10% NaCl. The precipitate was then dissolv¬ ed in guanidinium chloride (final concentration 6 M, pH 8.0) with 5% 2-mercaptoethanol and chromatographed on a Sephacryl S-300 or S-400 column, depending on the size of the adhesive precursor protein. The fractions containing the adhesive precursor protein were pooled, adjusted to pH 4.0 with glacial acetic acid, dialyzed against 5% acetic acid, then 0.1% acetic acid, with several changes of the dialysis solution, and lyophilized. The purified protein was characterized by its mobility on SDS-polyacrylamide gel electrophoresis using both Coumassie blue stain and Western blot analysis, its UV absorption spectrum, protein quanti- tation and amino acid composition analysis. EXAMPLE 21 The yeast cells were processed as in Example 20 up to the 10% NaCl precipitation step. Instead of treat¬ ing with NaCl, the supernatant was adjusted to pH 5.2 and centrifuged. The clear supernatant was dialyzed against 10% formic acid and slowly to water and then to 30 mM phosphate buffer (pH 7.3). After centrifuga¬ tion, the supernatant was chromatographed on a weak cation exchanger. The column was eluted first with a 0.2-1 M salt gradient at pH 7.3, followed by 1M NaCl in 5% acetic acid. The fractions containing the adhe¬ sive precursor protein were pooled, dialyzed and lyophilized as in Example 20, and the identity and purity of the adhesive precursor protein were deter¬ mined as describee in Example 20. EXAMPLE 22 After the yeast cells were broken and centrifuged as in Example 20, the pellet was solubilized in either 6 M guanidinium chloride or 8 M urea with 5% 2-mercap- toethenol. The insoluble cellular components were removed by centrifuging at 25,000 g for 30' at 4°C. The protein in the supernatant was precipitated by dialysis against water, and the adhesive precursor protein was extracted by a dilute formic acid or acetic acid solution (pH 2.0) for several hours. The adhesive precursor protein was precipitated by either adjusting the pH to neutral, or making the solution 10% NaCl. After centrifugation, the adhesive precur¬ sor protein pellet was dissolved in 6M guanidinium chloride and chromatographed on an S-300 or S-400 column depending on the size of the protein. The adhesive precursor protein, with the correct size, was pooled, adjusted to pH 4.0 with acetic acid, dialyzed against 5% acetic acid then 0.1% acetic acid and lyophilized. EXAMPLE 23 The yeast cells were broken and thoroughly washed as in Example 20. The pellet was extracted for several hours with dilute formic or acetic acid (pH 2.0). The solution was then adjusted to pH 3.0 and centrifuged. The adhesive precursor protein present in the supernatant was precipitated when the pH was adjusted to between 5.0 and 5.5. The adhesive precur¬ sor protein was collected by centrifugation, and, if necessary, further purified by either a weak cation exchange, or a gel filtration column as described in Examples 20 and 21.

Claims: CLAIMS What is claimed is: 1. A DNA segment comprising codons for the na¬ tive amino acid sequence of a bioadhesive precursor protein of a marine animal selected from the group consisting of mussels, barnacles and oysters. 2. A DNA segment as claimed in claim 1 wherein said marine animal is a mussel. 3. A DNA segment as claimed in claim 2 wherein said mussel is of the genus Mytilus . 4. A DNA segment as claimed in claim 3 , wherein the encoded bioadhesive precursor protein is of the mussel species Mytilus edulis. 5. A DNA segment as claimed . in claim 4 compris¬ ing codons for a bioadhesive precursor protein which contains the sequence : Lys Leu Ser Ser Tyr Lys Pro He Lys Thr Thr Tyr Asn Ala Lys Thr Asn Tyr Pro Pro Val Tyr Lys Pro Lys Met Thr Tyr Pro Pro Thr Tyr Lys Pro Lys Pro Ser Tyr Pro Pro Thr Tyr Lys Ser Lys Pro Thr Tyr Lys Pro Lys lie Thr Tyr Pro Pro Thr Tyr Lys Ala Lys Pro Ser Tyr Pro Pro Thr Tyr Lys Pro Lys Lys Thr Tyr Pro Pro Thr Tyr Lys Pro Lys Leu Thr Tyr Pro Pro Thr Tyr Lys Pro Lys Pro Ser Tyr Pro Pro Thr Tyr Lys Ser Lys Pro Thr Tyr Lys Pro Lys lie Thr Tyr Pro Pro Thr Tyr Lys Ala Lys Pro Ser Tyr Pro Pro Thr Tyr Lys Pro Lys Lys Thr Tyr Pro Pro Thr Tyr Lys Pro Lys Leu Thr Tyr Pro Pro Thr Tyr Lys Pro Lys Ala Ser Tyr Pro Pro Thr Tyr Lys Pro Lys Pro Ser Tyr Pro Pro Ser Tyr Lys Thr Lys Lys Thr Tyr Pro Pro Thr Tyr Lys Pro Lys Leu Thr Tyr Pro Pro Thr Tyr Lys Pro Lys Pro Ser Tyr Pro Pro Ser Tyr Lys Pro Lys Lys Thr Tyr CT/US87/03048 Pro Pro Thr Tyr Lys Pro Lys Leu Thr Tyr Pro Pro Thr Tyr Lys Ala Lys Pro Ser Tyr Pro Pro Thr Tyr Lys AU Lys Pro Ser Tyr Pro Pro Thr Tyr Lys Ala Lys Pre Ser Tyr Pro Pro Thr Tyr Lys Ala Lys Pro Str Tyr Pro Str Thr Tyr Lys Ala Lys Pro Ser Tyr Pro Pro Thr Tyr Lys Ala Lys Pro Thr Tyr Lys Ala Lys Pro Thr Tyr Lys AU Lys Pro Thr Tyr Pro Str Thr Tyr Lys AU Lys Pro Thr Tyr Pro Pre Thr Tyr Lys AU Lys Pre Ser Tyr Pro Pro Thr Tyr Lys AU Lys Pro Thr Tyr Lys AU Lys Pro Thr Tyr Lys AU Lys Pro Str Tyr Pro Pro Thr Tyr Lys AU Lys Pro Ser Tyr Pro Pro Thr Tyr Lys AU Lys Pro Ser Tyr Pro Pro Thr Tyr Lys AU Lys Pro Ser Tyr Pro Pro Thr Tyr Lys AU Lys Pro Ser Tyr Pro Pro Thr Tyr Lys Val Lys Pro Thr Tyr Lys A Lys Pre Thr Tyr Pro Ser Thr Tyr Lys AU Lys Pro Ser Tyr Pro Pre Thr Tyr Lys Ala Lys Pro Ser Tyr Pro Pro Thr Tyr Lys AU Lys Pre Ser Tyr Pro Pro Thr Tyr Lys Ala Lys Pro Ser Tyr Pre Pre Thr Tyr Lys AU Lys Pro Thr Tyr Pre Ser Thr Tyr Lys Ala Lys Pre Ser Tyr Pro Pro Thr Tyr Lys Pro Lys l e Ser Tyr Pro Pro Thr Tyr Lys Ala Lys Pro Ser Tyr Pre Ser Thr Tyr Lys Ala Lys Ser Ser Tyr Pro Pro Thr Tyr Lys Ala Lys Pro Ser Tyr Pre Pro Thr Tyr Lys Ala Lys Pre Thr Tyr Pre Ser Thr Tyr Lys Ala Lys Pro Thr Tyr Lys A Lys Pro Ser Tyr Pro Pro Thr Tyr Lys Ala Lys Pro Ser Tyr Pro Pro Thr Tyr Lys Ala Lys Pro Ser Tyr Pro Pro Thr Tyr Lys Ala Lys Pro Thr Tyr Lys Ala Lys Pro Thr Tyr Lys Ala Lys Pro Thr Tyr Pro Ser Thr Tyr Lys Ala Lys Pro Ser Tyr Pro Pre Thr Tyr Lys Ala Lys Pro Ser Tyr Pre Pre Thr Tyr Lys Ala Lys Pro Ser Tyr Pro Pro Thr Tyr Lys Ala Lys Pro Ser Tyr Pre Pre Thr Tyr Lys Ala Lys Pro Ser Tyr Pro Pro Thr Tyr Lys Ala Lys Pro Ser Tyr Pro Pro Thr Tyr Lys AU Lys Pre Ser Tyr Pro Pre Thr Tyr Lys Ala Lys Pro Thr Tyr Lys AU Lys Pro Thr Tyr Pro Ser Thr Tyr Lys Ala Lys Pro Ser Tyr Pro Pro Thr Tyr Lys Ala Lys Pro Ser Tyr Pro Pro Thr Tyr Lys Ala Lys Pro Ser Tyr Pro Pro Thr Tyr Lys AU Lys Pro Thr Tyr Lys Ala Lys Pro Thr Tyr Pro Ser Thr Tyr Lys AU Lys Pro Ser Tyr Pro Pro Thr His Lys Ala Lys Pro Thr Tyr Lys AU Lys Pro Thr Tyr Pre Ser Thr Tyr Lys Ala Lys Pre Thr Tyr Pro Ser Thr Asp Gly Ala Lys Ser, or a fragment there©!. -69- 6. A DNA segment as claimed in claim 4, coding for a bioadhesive precursor protein having the amino acid sequence: Lys Pro Lys Pro 8er Tyr Pro Pro Str Tyr Lys Pro Lys Thr Thr Tyr Pro Pro Thr Tyr Lys Pro Lys He Ser Tyr Pro Pro Thr Tyr Lys Ala Lys Pro Ser Tyr Pro Ala Thr Tyr Lys Ala Lys Pro Ser Tyr Pro Pro Thr Tyr Lys Ala Lys Pro Ser Tyr Pro Pro Thr Tyr Lys Ala Lys Pro Ser Tyr Pro Pro Thr Tyr Lys Ala Lys Pro Thr Tyr Lys Ala Lys Pro Thr Tyr Pro Pro Thr Tyr Lys Ala Lye Pro Ser Tyr Pro Pro Thr Tyr Lys Pro Lys Pro Ser Tyr Pro Pro Thr Tyr Lys Ser Lys Ser He Tyr Pro Ser Ser Tyr Lys Pro Lys Lys Thr Tyr Pro Pro Thr Tyr Lys Pro Lys Leu Thr Tyr Pro Pro Thr Tyr Lys Pro Lys Pro Ser Tyr Pro Pro Ser Tyr Lys Pro Lys He Thr Tyr Pro Ser Thr Tyr Lys Leu Lys Pro Ser Tyr Pro Pro Thr Tyr Lys Ser Lys Thr Ser Tyr Pro Pro Thr Tyr Asn Lys Lys He Ser Tyr Pro Ser Ser Tyr Lys Ala Lys Thr Ser Tyr Pro Pro Ala Tyr Lys Pro Thr Asn Arg Tyr or a fragment thereof. 87/03048 -70- 7. A DNA segment as claimed in claim 4, coding for a bioadhesive precursor protein having the amino acid sequences L u Pro Pro Ala Pro Pro Ala Pro Ala Phe Ala Pro Ala Pro Ala Leu Ala Pro Ala Pro Pro Asn Pro Asn Pro Pro Ser Pro Pro Ser Pro Pro Ser Pro Pro Thr f o Pro Pro Thr Pro Pro Ser Pro Pro Ala Pro Pro Ser Pro Pro Pro Ser Pro Pro Asn Pro Pro Pro Ser Pro Pro Pro Ser Pro Pro Phe Pro Pro Ala Pro Pro Pro Ser Pro Pro Phe Pro Pro Thr Tyr Lys Ala Lys Pro Thr Tyr Lys Ala Lys Pro Thr Asn Pro Ser Thr Tyr Lys Ala Lys Pro Ser Tyr Pro Pro Thr Tyr Lys Ala Lys Pro Ser Tyr Pro Pro Thr Tyr Lys Ala Lys Pro Ser Tyr Pro Pro Thr Tyr Lys Ala Lys Pro Thr Tyr Lys He Lys Pro Thr Tyr Pro Ser Thr Tyr Lys Ala Lys Pro Thr Asn Pro Ser Thr Tyr Lys Ala Lys Pro Ser Tyr Pro Pro Thr Tyr Lys Ala Lys Pro Ser Tyr Pro Pro Thr Tyr Lys Ala Lys Pro Ser Tyr Pro Pro Thr Tyr Lys Ala Lys Pro Thr Tyr Lys He Lys Pro Thr Tyr Pro Ser Thr Tyr Lys Ala Lys Pro Th Tyr Lys Ala Lys Pro Thr Tyr Pro Pro Thr Tyr Lys Ala Lys Pro Ser. Tyr Pro Pro Thr Tyr Lys Pro Lys Pro Ser Tyr Pro Pro Thr Tyr Lys Ser Lys Ser He Tyr Pro Ser Ser Tyr Lys Pro Lys Lys Thr Tyr Pro Pro Thr Tyr Lys Pro Lys Leu Thr Tyr Pro Pro Thr Tyr Lys Pro Lys Pro Ser Tyr Pro Pro Ser Tyr Lys Pro Lys He Thr Tyr Pro Ser Thr Tyr Lys Leu Lys Pro Ser Tyr Pro Pro Thr Tyr Lys Ser Lys Thr Ser Tyr Pro Pro Thr Tyr Asn Lys Lys He Ser Tyr Pro Ser Ser Tyr Lys Ala Lys Thr Ser Tyr Pro Pro Ala Tyr Lys Pro Thr Asn Arg Tyr or a fragment thereof. -71- β. A DNA segment as claimed in claim 4, coding for a bioadhesive precursor protein having the amino acid sequence: Tyr Lys Ala Lys Pro S*t Tyr Pro Pro Thr Tyr Lys Ala Lys Pro Thr Tyr Lys Ala Lys Pro Thr Asn Pro Ser Thr Tyr Lys Ala Lys Pro Ser Tyr Pro Pro Thr Tyr Lys Ala Lys Pro Ser Tyr Pro Pro Thr Tyr Lys Ala Lys Pro Ser Tyr Pro Pro Thr Tyr Lys Ala Lys Pro Thr Tyr Lys Ala Lys Pro Thr Tyr Pro Ser Thr Tyr Lys Ala Lys Pro Thr Tyr Lys Ala Lys Pro Thr Tyr Pro Pro Thr Tyr Lys Ala Lys Pro Ser Tyr Pro Pro Thr Tyr Lys Pro Lys Pro Ser Tyr Pro Pro Thr Tyr Lys Ser Lys Ser He Tyr Pro Ser Ser Tyr Lys Pro Lys Lys Thr Tyr Pro Pro Thr Tyr, or a fragment thereof. 9. A DNA segment as claimed in claim 4, coding for a bioadhesive precursor protein having the amino acid sequence: Thr Ser Thr Tyr Lys Ala Lys Pro Ser Tyr Pro Pro Thr Tyr Lys Ala Lys Pro Thr Tyr Pro Ser Thr Tyr Lys Ala Lys Pro Ser Tyr Pro Pro Thr Tyr Lys Pro Lys He Ser Tyr Pro Pro Thr Tyr Lys Ala Lys Pro Ser Tyr Pro Pro Thr Tyr Lys Ala Lys Pro Ser Tyr Pro Pro Thr Tyr Lys Ala Lys Pro Thr Tyr Lys Ala Lys Pro Thr Asn Pro Ser Thr Tyr Lys Ala Lys Pro Ser Tyr, or a fragment thereof. -72- 10. A DNA segment as claimed in claim 4, compris¬ ing the following DNA sequence: AAG CT6 TCA TCT TAC AAA CCT ATT AA6 ACA ACA TAT AAT GCA AAG ACA AAT TAT CCA CCA STT TAT AAA CCT AAG ATS ACT TAT CCT CCT ACA TACAAA CCA AAG CCC AGT TAT CCT CCA ACA TAT AAA TCA AAG CCC ACA TAC AAA CCT AAG ATA ACA TAC CCT CCA ACT TAT AAA 6CA AAG CCC AGT TAT CCT CCA ACA TAT AAA CCT AAG AAA ACT TAT CCC CCC ACA TAT AAA CCT AAA CTA ACC TAT CCT CCT ACA TAC AAA CCA AAG CCC AGT TAT CCT CCA ACA TAT AAA TCA AAG CCC ACA TAC AAA CCT AAG ATA ACA TAC CCT CCA ACA TAT AAA GCA AAG CCA AGT TAT CCT CCT ACA TAT AAA CCT AAG AAA ACT TAT CCC CCC ACA TAT AAA CCT AAA CTA ACC TAT CCT CCT ACA TAT AAA CCA AAG GCC AGT TAT CCT CCA ACA TAT AAA CCA AAA CCA AGT TAT CCC CCT TCA TAT AAA ACT AAG AAA ACT TAT CCC CCC ACA TAT AAA CCT AAA CTA ACC TAT CCT CCT ACA TAT AAA CCA AAA CCA AGT TAT CCC CCT TCA TAC AAA CCT AAG AAA ACT TAT CCC CCC ACA TAT AAA CCT AAA CTA ACC TAT CCT CCT ACA TAT AAA GCA AAG CCA AGT TAT CCT CCA ACA TAT AAA GCA AAA CCA AGT TAT CCT CCA ACT TAT AAA GCA AAA CCA AGT TAT CCT CCA ACA TAT AAA GCA AAA CCA AGT TAT CCT TCA ACT TAT AAA GCA AAA CCA AGT TAT CCT CCA ACT TAT AAA GCA AAG CCA ACT TAT AAA GCA AAG CCA ACT TAT AAA GCA AAG CCA ACT TAT CCT TCA ACG TAT AAA GCA AAG CCA ACT TAT CCT CCA ACT TAT AAA GCA AAA CCA AGT TAT CCT CCA ACT TAT AAA GCA AAG CCA ACT TAT AAA GCA AAG CCA ACT TAT AAA GCA AAG CCA AGT TAT CCT CCA ACT TAT AAA GCA AAA CCA AGT TAT CCT CCA ACT TAT AAA GCA AAG CCA AGT TAT CCT CCA ACG TAT AAA GCA AAG CCA AGT TAT CCT CCA ACT TAT AAA GCA AAA CCA AGT TAT CCT CCA ACT TAT AAA 6TA AAG CCA ACA TAT AAA GCA AAA CCA ACT TAT CCT TCA ACG TAT AAA GCA AAG CCA AGT TAT CCT CCA ACT TAT AAA GCA AAA CCA A6T TAT CCT CCAACT TAT AAA GCA AAA CCA AGT TAT CCT CCA AC6 TAT AAA GCA AAA CCA AGT TAT CCT CCA ACT TAT AAA GCA AAA CCA ACT TAT CCT TCA ACG TAT AAA GCA AAA CCA AGT TAT CCT CCA ACT TAT AAA CCT AAA ATA AGT TAT CCT CCA ACT TAT AAA GCA AAA CCA AGT TAT CCT TCA ACT TAT AAA GCA AAA TCA AGT TAT CCT CCA ACT TAT AAA GCA AAA CCA AGT TAT CCT CCA ACT TAT AAA GCA AAG CCA ACT TAT CCT TCA ACG TAT AAA GCA AAG CCA ACT TAT AAA GCA AAG CCA AGT TAT CCT CCA ACG TAT AAA GCA AAG CCA AGT TAT CCT CCA ACT TAT AAA GCA AAA CCC AGT TAT CCT CCA ACT TAT AAA GCA AAG CCA ACT TAT AAA GCA AAG CCA ACT TAT AAA GCA AAG CCA ACT TAT CCT TCA ACG TAT AAA GCA AAG CCA AGT TAT CCT CCA ACT TAT AAA GCA AAACCA AGT TAT CCT CCA ACT TAT AAA GCA AAA CCA AGT TAT CCT CCA ACT TAT AAA GCA AAA CCA AGT TAT CCT CCA ACT TAT AAA GCA AAG CCA AGT TAT CCT CCA ACG TAT AAA GCA AAG CCA AGT TAT CCT CCA ACT TAT AAA GCA AAA CCA AGT TAT CCT CCA ACT TAT AAA GCA AAG CCA ACT TAT AAA GCA AAG CCA ACT TAT CCT TCA ACG TAT AAA GCA AAG CCA AGT TAT CCT CCA ACT TAT AAA GCA AAA tCA AGT TAT CCT CCA ACT TAT AAA GCA AAA CCA AGT TAT CCT CCA ACT TAT AAA GCA AAA CCA ACT TAT AAA GCA AAG CCA ACT TAT CCT TCA ACG TAT AAA GCA AAA CCA AGT TAT CCT CCA ACT CAT AAA GCA AAA CCA ACT TAT AAA GCA AAG CCA ACT TAT CCT TCA ACT TAT AM GCA AAA CCA ACT TAT CCT TCA ACG GAT 6GA GCA AM TCA. or a fragment thereof. 11. A DNA segment ss claimed in claim 4, compris¬ ing the following DNA sequence: AAA CCA AAA CCA AGT TAT CCA CCA TCT TAT AAA CCT AAA ACA ACT TAT CCT CCA ACT TAT AAA CCT AAG ATA AGT TAT CCT CCA ACT TAT AAA GCA AAA CCA AGT TAT CCA GCA ACT TAT AAA GCA AAA CCA AGT TAT CCT CCA ACT TAT AAA GCA AAA CCA AGT TAT CCT CCA ACT TAT AAA GCA AAA CCA AGT TAT CCT CCA ACT TAT AAA GCA AAG CCA ACT TAT AAA GCA AAG CCA ACT TAT CCT CCA ACT TAT AAA GCA AAA CCA AGT TAT CCT CCA ACA TAT AAA CCA AAG CCA AGT TAT CCT CCA ACT-TAT AAA TCC AAG TCA ATA T T CCC TCT TCA TAC AAA CCT AAG AAA ACT TAT CCC CCC ACA TAT AAA CCT AAA CTA ACC TAT CCT CCA ACA TAT AAA CCA AAG CCA AGT TAT CCA CCA TCT TAT AAA CCT AAG ATT ACT TAT CCC TCA ACT TAT AAA TTG AAG CCA AGT TAT CCT CCA ACA TAC AAA TCT AAA ACA AGT TAC CCT CCT ACA TAT AAC AAA AAG ATC AGC TAT CCA TCA TCA TAT AAA GCT AAG ACA AGT TAT CCC CCA GCA TAT AAA CCA ACA AAC AGA TAT, or a fragment thereof. -75- 12. A DNA segment as claimed in claim 4, compris¬ ing the following DNA sequence: CTA CCG CCA GCA CCT CCT GCT CCT GCA TTT GCT CCT GCT CCT GCA CTT GCA CCG GCT CCT CCA AAT CCT AAT CCT CCA AGT CCT CCG AGT CCT CCG AGT CCA CCG ACT CCA CCG CCG ACT CCT CCA AGT CCA CCA GCT CCT CCG AGT CCA CCA CCG AGT CCT CCG AAT CCA CCT CCG AGT CCA CCT CCG AGT CCA CCG TTT CCT CCG GCT CCA CCT CCG AGT CCA CCA TTT CCT CCG ACT TAT AAA GCA AAG CCA ACT TAT AAA GCA AAG CCA ACT AAT CCT TCA ACG TAT AAA GCA AAG CCA AGT TAT CCT CCA ACT TAT AAA GCA AAA CCA AGT TAT CCT CCA ACT TAT AAA GCA AAA CCA AGT TAT CCT CCA ACG TAT AAA GCA AAG CCA ACT TAT AAA ATA AAG CCA ACT TAT CCT TCA ACG TAT AAA GCA AAG CCA ACT AAT CCT TCA ACG TAT AAA GCA AAG CCA AGT TAT CCT CCA ACT TAT AAA GCA AAA CCA AGT TAT CCT CCA ACT TAT AAA GCA AAA CCA AGT TAT CCT CCA ACG TXT AAA GCA AAG CCA ACT TAT AAA ATA AAG CCA ACT TAT CCT TCA ACG TAT AAA GCA AAG CCA ACT TAT AAA GCA AAG CCA ACT TAT CCT CCA ACT TAT AAA GCA AAA CCA AGT TAT CCT CCA ACA TAT AAA CCA AAG CCA AGT TAT CCT CCA ACT TAT AAA TCC AAG TC*. ATA TAT CCC TCT TCA TAC AAA CCT AAG AAA ACT TAT CCC CCC ACA TAT AAA CCT AAA CTA ACC TAT CCT CCA ACA TAT AAA CCA AAG CCA AGT TAT CCA CCA TCT TAT AAA CCT AAG ATT ACT TAT CCC TCA ACT TAT AAA TTG AAG CCA AGT TAT CCT CCA ACA TAC AAA TCT AAA ACA AGT TAC CCT CCT ACA TAT AAC AAA AAG ATC AGC TAT CCA TCA TCA TAT AAA GCT AAG ACA AGT TAT CCC CCA GCA TAT AAA CCA ACA AAC AGA TAT, or a fragment thereof. -76- 13. A DNA segment as claimed in claim 4 compris¬ ing the following DNA sequence: TAT AAA GCA AAA CCA AGT TAT CCT CCA ACT TAT AAA GCA AAG CCA ACT TAT AAA GCA AAG CCA ACT AAT CCT TCA ACG TAT AAA GCA AAG CCA AGT TAT CCT CCA ACT TAT AAA GCA AAA CCA AGT TAT CCT CCA ACT TAT AAA GCA AAA CCA AGT TAT CCT CCA ACG TAT AAA GCA AAG CCA ACT TAT AAA GCA AAG CCA ACT TAT CCT TCA ACG TAT AAA GCA AAG CCA ACT TAT AAA GCA AAG CCA ACT TAT CCT CCA ACT TAT AAA GCA AAA CCA AGT TAT CCT CCA ACA TAT AAA CCA AAG CCA AGT TAT CCT CCA ACT TAT AAA TCC AAG TCA ATA TAT CCC TCT TCA TAC AAA CCT AAG AAA ACT TAT CCC CCC ACA TAT, or a fragment thereof. -77- 14. A DNA segment as claimed in claim 4 compris¬ ing the following DNA sequence: ACT TCA ACT TAT AAA GCA AAA CCA AGT TAT CCT CCA ACT TAT AAA GCA AAG CCA ACT TAC CCT TCA ACG TAT AAA GCA AAA CCA AGT TAT CCT CCA ACT TAT AAA CCT AAG ATA AGT TAT CCT CCA ACT TAT AAA GCA AAA CCA AGT TAT CCT CCA ACT TAT AAA GCA AAA CCA AGT TAT CCT CCA ACT TAT AAA GCA AAG CCA ACT TAT AAA GCA AAG CCA ACT AAT CCT TCA ACG TAT AAA GCA AAG CCA AGT TAT, or a fragment thereof: 15. A vector for expression of a bioadhesive pre¬ cursor protein comprising: (a) a DNA segment comprising codons for the native amino acid sequence of a bioad¬ hesive precursor protein of a marine animal selected from the group consist¬ ing of mussels, barnacles, and oysters. (b) a promoter and transcription initiation signals, operably linked to said DNA segment, which are capable of effecting microbial expression of the bioadhesive precursor protein. 16. A vector as claimed in claim IS wherein the encoded bioadhesive precursor protein is from mussel. 17. A vector as claimed in claim 16, wherein the encoded bioadhesive precursor protein is of the mussel species Mytilus edulis. 18. A vector as claimed in claim 17, wherein the DNA segment (a) comprises codons for a bioadhesive precursor comprising the amino acid sequence! Lys Ltu Ser Ser Tyr Lys Pro lie Lys Thr Thr Tyr Asn AU Lys Thr Asn Tyr Pro Pro Val Tyr Lys Pre Lys Met Thr Tyr Pro Pro Thr Tyr Lys Pro Lys Pro Str Tyr Pro Pro Thr Tyr Lys Ser Lys Pre Thr Tyr Lys Pro Lys lie Thr Tyr Pre Pre Thr Tyr Lys AU Lys Pro Ser Tyr Pro Pro Thr Tyr Lys Pro Lys Lys Thr Tyr Pro Pro Thr Tyr Lys Pre Lys Leu Thr Tyr Pro Pro Thr Tyr Lys Pro Lys Pro Ser Tyr Pro Pro Thr Tyr Lys Ser Lys Pro Thr Tyr Lys Pro Lys lit Thr Tyr Pro Pro Thr Tyr Lys AU Lys Pro Ser Tyr Pro Pro Thr Tyr Lys Pre Lys Lys Thr Tyr Pro Pro Thr Tyr Lys Pro Lys Leu Thr Tyr Pro Pro Thr Tyr Lys Pro Lys AU Ser Tyr Pro Pro Thr Tyr Lys Pro Lys Pro Ser Tyr Pro Pro Ser Tyr Lys Thr Lys Lys Thr Tyr Pro Pre Thr Tyr Lys Pro Lys Leu Thr Tyr Pro Pro Thr Tyr Lys Pre Lys Pro Ser Tyr Pre Pro Ser Tyr Lys Pro Lys Lys Thr Tyr Pro Pro Thr Tyr Lys Pro Lys Leu Thr Tyr Pro Pro Thr Tyr Lys Ala Lys Pro Ser Tyr Pro Pro Thr Tyr Lys AU Lys Pro Ser Tyr Pro Pro Thr Tyr Lys Ala Lys Pro Ser Tyr Pro Pro Thr Tyr Lys A . Lys. Pre Ser Tyr Pro Ser Thr Tyr Lys Ala Lys Pro Ser Tyr Pro Pro Thr Tyr Lys Ala Lys Pro Thr Tyr Lys Ala Lys Pro Thr Tyr Lys AU Lys Pro Thr Tyr Pro Ser Thr Tyr Lys Ala -Lys Pro Thr- Tyr. Pro Pro Thr Tyr Lys Ala Lys Pro Ser Tyr Pro Pro Thr Tyr Lys Ala Lys Pro Thr Tyr Lys Ala Lys Pro Thr Tyr Lys Ala Lys Pre Ser Tyr Pre Pre Thr Tyr Lys Ala Lys Pre Ser Tyr Pre Pro Thr Tyr Lys Ala Lys Pro Ser Tyr Pro Pre Thr Tyr Lys Ala Lys Pre Ser Tyr Pro Pro Thr Tyr Lys Ala Lys Pre Ser Tyr Pro Pro Thr Tyr Lys Val Lys Pro Thr Tyr Lys AU Lys Pro Thr Tyr Pro Ser Thr Tyr Lys AU Lys Pro Ser Tyr Pro Pro Thr Tyr Lys Ala Lys Pro Ser Tyr Pro Pro Thr Tyr Lys AU Lys Pro Ser Tyr Pro Pro Thr Tyr Lys Ala Lys Pro Ser Tyr Pro Pro Thr Tyr Lys AU Lys Pro Thr Tyr Pro Ser Thr Tyr Lys Ala Lys Pro Ser Tyr Pro Pre Thr Tyr Lys Pro Lys l e Ser Tyr Pro Pre Thr Tyr Lys AU Lys Pro Ser Tyr Pre Ser Thr Tyr Lys Ala Lys Ser Ser Tyr Pre Pre Thr Tyr Lys Ala Lys Pre Ser Tyr Pro Pro Thr Tyr Lys Ala Lys Pro Thr Tyr Pre Ser Thr Tyr Lys Ala Lys Pro Thr Tyr Lys A Lys Pro Ser Tyr Pro Pro Thr Tyr Lys Ala Lys Pro Ser Tyr Pro Pro Thr Tyr -79- Lys AU Lys Pro Str Tyr Pro Pro Thr Tyr Lys AU Lys Pro Thr Tyr Lys AU Lys P Thr Tyr Lys AU Lys Pro Thr Tyr Pro Ser Thr Tyr Lys Ala Lys Pro Ser Tyr Pro P Thr Tyr Lys Ala Lys Pro Str Tyr Pro Pro Thr Tyr Lys Ala Lys Pro Str Tyr Pro P Thr Tyr Lys Ala Lys Pro Str Tyr Pro Pro Thr Tyr ly% Ala Lys Pro Ser Tyr Pro P Thr Tyr Lys A Lys Pro Str Tyr Pro Pro Thr Tyr Lys AU Lys Pre Ser Tyr Pro P Thr Tyr Lys Ala Lys Pro Thr Tyr Lys AU Lys Pro Thr Tyr Pro Str Thr Tyr Lys Al Lys Pro Str Tyr Pro Pro Thr Tyr Lys AU Lys Pro Ser Tyr Pro Pro Thr Tyr Lys A Lys Pro Ser Tyr Pro Pro Thr Tyr Lys AU Lys Pro Thr Tyr Lys Ala Lys Pro Thr Ty Pro Ser Thr Tyr Lys A Lys Pro Ser Tyr Pro Pro Thr His Lys Ala Lys Pro T h r Ty Lys Ala Lys Pro Thr Tyr Pro Ser Thr Tyr Lys Ala Lys Pro Thr Tyr Pro Ser T h r As Gly Ala Lys Ser, or a fragment thereof. 19. A vector as claimed in claim 17, wherein the DNA segment ( a ) comprises codons for a bioadhesive precursor comprising the amino acid.sequence: Lys Pro Lys Pro Ser Tyr Pro Pro Ser Tyr Lys Pro Lys Thr Thr Tyr Pro Pro Thr Tyr Lys Pro Lys He Ser Tyr Pro Pro Thr Tyr Lys Ala Lys Pro Ser Tyr Pro Ala Thr Tyr Lys Ala Lys Pro Ser Tyr Pro Pro Thr Tyr Lys Ala Lys Pro Ser Tyr Pro Pro Thr Tyr Lys Ala Lys Pro Ser Tyr Pro Pro Thr Tyr Lys Ala Lys Pro Thr Tyr Lys Ala Lys Pro Thr Tyr Pro Pro Thr Tyr Lys Ala Lys Pro Ser Tyr Pro Pro Thr Tyr Lys Pro Lys Pro Ser Tyr Pro Pro Thr Tyr Lys Ser Lys Ser He Tyr Pro Ser Ser Tyr Lys Pro Lys Lys Thr Tyr Pro Pro Thr Tyr Lys Pro Lys Leu Thr Tyr Pro Pro Thr Tyr Lys Pro Lys Pro Ser Tyr Pro Pro Ser Tyr Lys Pro Lys He Thr Tyr Pro Ser Thr Tyr Lys Leu Lys Pro Ser Tyr Pro Pro Thr Tyr Lys Ser Lys Thr Ser Tyr Pro Pro Thr Tyr Asn Lys Lys -80- Zle Ser Tyr Pro Ser Ser Tyr Lys Ala Lys Thr Ser Tyr Pro Pro Ala Tyr Lys Pro Thr Asn Arg Tyr or a fragment thereof. 20. A vector as claimed in claim 17, wherein the DNA segment (a) comprises codons for a bioadhesive precursor protein comprising the amino acid sequence: Leu Pro Pro Ala Pro Pro Ala Pro Ala Phe Ala Pro Ala Pro Ala Leu Ala Pro Ala Pro Pro Asn Pro Asn Pro Pro Ser Pro Pro Ser Pre Pro Ser Pro Pro Thr Pro Pro Pro Thr Pro Pro Ser Pro Pre Ala Pro Pre Ser Pro Pre Pre Ser Pro Pre Asn Pro Pro Pro Ser Pro Pro Pro Ser Pro Pro Phe Pro Pro Ala Pro Pro Pro Ser Pro Pro Phe Pro Pro Thr Tyr Lys Ala Lys Pro Thr Tyr Lys Ala Lys Pro Thr Asn Pro Ser Thr Tyr Lys Ala Lys Pro Ser Tyr Pro Pro Thr Tyr Lys Ala Lys Pro Ser Tyr Pro Pro Thr Tyr Lys Ala Lys Pro Ser Tyr Pro Pro Thr Tyr Lys Ala Lys Pro Thr Tyr Lys He Lys Pro Thr Tyr Pro Ser Thr Tyr Lys Ala Lys Pro Thr Asn Pro Ser Thr Tyr Lys Ala Lys Pro Ser Tyr Pro Pro Thr Tyr Lys Ala Lys Pro Ser Tyr Pro Pro Thr Tyr Lys Ala Lys Pro Ser Tyr Pro Pro Thr Tyr Lys Ala Lys Pro Thr Tyr Lys He Lys Pro Thr Tyr Pro Ser Thr Tyr Lys Ala Lys Pro Thr Tyr Lys Ala Lys Pro Thr Tyr Pro Pro Thr Tyr Lys Ala Lys Pro Ser Tyr Pro Pro Thr Tyr Lys Pro Lys Pro Ser Tyr Pro Pro Thr Tyr Lys Ser Lys Ser He Tyr Pro Ser Ser Tyr Lys Pro Lys Lys Thr Tyr Pro Pro Thr Tyr Lys Pro Lys Leu Thr Tyr Pro Pro Thr Tyr Lys Pro Lys Pro Ser Tyr Pro Pro Ser Tyr Lys Pro Lys He Thr Tyr Pro Ser Thr Tyr Lys Leu Lys Pro Ser Tyr Pro Pro Thr Tyr Lys Ser Lys Thr Ser Tyr Pro Pro Thr Tyr Asn Lys Lys He Ser Tyr Pro Ser Ser Tyr Lys Ala Lys Thr Ser Tyr Pro Pro Ala Tyr Lys Pro Thr Asn Arg Tyr or a fragment thereof. -81- 21. A vector M claimed in claim 17 wherein the DNA segment (s) comprises codons for a bioadhesive precursor protein comprising the amino acid sequence: Tyr Lys Ala Lys Pro Ser Tyr Pro Pro Thr Tyr Lys Ala Lys Pro Thr Tyr Lys Ala Lys Pro Thr Asn Pro Sen Thr Tyr Lys Ala Lys pro Ser Tyr Pro Pro Thr Tyr Lys Ala Lys ro B it Tyr Pro Pro Thr Tyr Lys Ala Lys Pro Ser Tyr Pro Pro Thr Tyr Lys Ala Lys Pro Thr Tyr Lys Ala Lys Pro Thr Tyr Pro Ser Thr Tyr Lys Ala Lys Pro Thr Tyr Lys Ala Lys Pro Thr Tyr Pro Pro Thr Tyr Lys Ala Lys Pro Ser Tyr Pro Pro Thr Tyr Lys Pro Lys Pro Ser Tyr Pro Pro Thr Tyr Lys Ser Lys Ser He Tyr Pro Ser Ser Tyr Lys Pro Lys Lys Thr Tyr Pro Pro Thr Tyr, or a fragment thereof. 22. A vector as claimed in claim 17 wherein the DNA segment (a) comprises codons for a bioadhesive precursor protein comprising the amino acid sequence: Thr Ser Thr Tyr Lys Ala Lys Pro Ser Tyr Pro Pro Thr Tyr Lys Ala Lys Pro Thr Tyr Pro Ser Thr Tyr Lys Ala Lys Pro Ser Tyr Pro Pro Thr Tyr Lys Pro Lys He Ser Tyr Pro Pro Thr Tyr Lys Ala Lys Pro Ser Tyr Pro Pro Thr Tyr Lys Ala Lys Pro Ser Tyr Pro Pro Thr Tyr Lys Ala Lys Pro Thr Tyr Lys Ala Lys Pro Thr Asn Pro Ser Thr Tyr Lys Ala Lys Pro Ser Tyr, or a fragment thereof. -82- 23• A vector as claimed in claim 17, wherein the DNA segment ( a ) consists of the following sequence of bases: MG CTG TCA TCT TAC AM CCT ATT MG ACA ACA TAT MT GCA MG ACA MT TAT CCA CCA GTT TAT AM CCT MG ATG ACT TAT CCT CCT ACA TAC AM CCA MG CCC AGT TAT CCT CCA ACA TAT AM TCA MG CCC ACA TAC AM CCT MG ATA ACA TAC CCT CCA ACT TAT AM GCA MG CCC AGT TAT CCT CCA ACA TAT MA CCT MG MA ACT TAT CCC CCC ACA TAT MA CCT MA CTA ACC TAT CCT CCT ACA TAC AM CCA MG CCC AGT TAT CCT CCA ACA TAT MA TCA MG CCC ACA TAC MA CCT MG ATA ACA TAC CCT CCA ACA TAT AM GCA AAG CCA AGT TAT CCT CCT ACA TAT AM CCT MG MA ACT TAT CCC CCC ACA TAT AM CCT AM CTA ACC TAT CCT CCT ACA TAT MA CCA KAQ GCC AGT TAT CCT CCA ACA TAT MA CCA AM CCA AGT TAT CCC CCT TCA TAT AM ACT MG MA ACT TAT CCC CCC ACA TAT AM CCT AM CTA ACC TAT CCT CCT ACA TAT AM CCA AM CCA AGT TAT CCC CCT TCA TAC AM CCT MG AM ACT TAT CCC CCC ACA TAT MA CCT MA CTA ACC TAT CCT CCT ACA TAT AM GCA AAG CCA AGT TAT CCT CCA ACA TAT AM GCA AM CCA AGT TAT CCT CCA ACT TAT AM GCA AM CCA AGT TAT CCT CCA ACA TAT AM GCA AM CCA A6T TAT CCT TCA ACT TAT AM GCA AM CCA AGT TAT CCT CCA ACT TAT AM GCA MG CCA ACT TAT AM GCA MG CCA ACT TAT AM GCA MG CCA ACT TAT CCT TCA ACG TAT AM GCA MG CCA ACT TAT CCT CCA ACT TAT AM GCA MA CCA AGT TAT CCT CtA ACT TAT AM GCA MG CCA ACT TAT AM GCA MG CCA ACT TAT AM GCA AAG CCA AGT TAT CCT CCA ACT TAT AM GCA MA CCA AGT TAT CCT CCA ACT TAT AAA CA -83- MG CCA AGT TAT CCi' CCA ACG TAT AM 6CA MG CCA AGT TAT CCT CCA ACT TAT AM GCA AM CCA AGT TAT CCT CCA ACT TAT AM GTA MG CCA ACA TAT AM GCA AM CCA ACT TAT CCT TCA ACG TAT AM GCA MG CCA AGT TAT CCT CCA ACT TAT MA GCA AM CCA AGT TAT CCT CCA ACT TAT MA SCA AM CCA AGT TAT CCT CCA ACG TAT AM GCA AM CCA AGT TAT CCT CCA ACT TAT AM GCA AM CCA ACT TAT CCT TCA ACG TAT AM GCA AM CCA AGT TAT CCT CCA ACT TAT MA CCT AM ATA AGT TAT CCT CCA ACT TAT AM GCA AM CCA AGT TAT CCT TCA ACT TAT MA GCA MA TCA AGT TAT CCT CCA ACT TAT AM GCA AAA CCA AGT TAT CCT CCA ACT TAT MA GCA MG CCA ACT TAT CCT TCA ACG TAT MA GCA MG CCA ACT TAT MA GCA MG CCA AGT TAT CCT CCA ACG TAT AM GCA MG CCA AGT TAT CCT CCA ACT TAT MA GCA MA CCC AGT TAT CCT CCA ACT TAT MA GCA MG CCA ACT TAT MA GCA MG CCA ACT TAT AM GCA MG CCA ACT TAT CCT TCA ACG TAT AM GCA MG CCA AGT TAT CCT CCA ACT TAT AM GCA AM CCA AGT TAT CCT CCA ACT TAT AM GCA AM CCA AGT TAT CCT CCA ACT TAT MA GCA AAA CCA AGT TAT CCT CCA ACT TAT AM GCA MG CCA AGT TAT CCT CCA ACG TAT AM GCA MG CCA AGT TAT CCT CCA ACT TAT AM GCA AM CCA AGT TAT CCT CCA ACT TAT AM GCA MG CCA ACT TAT AM GCA MG CCA ACT TAT CCT TCA ACG TAT AM GCA MG CCA AGT TAT CCT CCA ACT TAT MA GCA AM «A AGT TAT CCT CCA ACT TAT AM GCA MA CCA AGT TAT CCT CCA ACT TAT AM GCA AM CCA ACT TAT AM GCA MG CCA ACT TAT -84- CCT TCA ACG TAT AM GCA AM CCA AGT TAT CCT CCA ACT CAT AM GCA AM CCA ACT TAT AM GCA MG CCA ACT TAT CCT TCA ACT TAT AM GCA AM CCA ACT TAT CCT TCA ACG GAT GGA GCA AM TCA. or a fragment thereof. 24. A vector as claimed in claim 17, wherein the DNA segment (a) consists of the following sequence of bases: AAA CCA AAA CCA AGT TAT CCA CCA TCT TAT AAA CCT AAA ACA ACT TAT CCT CCA ACT TAT AAA CCT AAG ATA AGT TAT CCT CCA ACT TAT AAA GCA AAA CCA AGT TAT CCA GCA ACT TAT AAA GCA AAA CCA AGT TAT CCT CCA ACT TAT AAA GCA AAA CCA AGT TAT CCT CCA ACT TAT AAA GCA AAA CCA AGT TAT CCT CCA ACT TAT AAA GCA AAG CCA ACT TAT AAA GCA AAG CCA ACT TAT CCT CCA ACT TAT AAA GCA AAA CCA AGT TAT CCT CCA ACA TAT AAA CCA AAG CCA AGT TAT CCT CCA ACT TAT AAA TCC AλG TCA ATA TAT CCC TCT TCA TAC AAA CCT AAG AAA ACT TAT CCC CCC ACA TAT AAA CCT AAA CTA ACC TAT CCT CCA ACA TAT AAA CCA AAG CCA AGT TAT CCA CCA TCT TAT AAA CCT AAG ATT ACT TAT CCC TCA ACT TAT AAA TTG AAG CCA AGT TAT CCT CCA ACA TAC AAA TCT AAA ACA AGT TAC CCT CCT ACA TAT AAC AAA AAG ATC AGC TAT CCA TCA TCA TAT AAA GCT AAG ACA AGT TAT CCC CCA GCA TAT AAA CCA ACA AAC AG TAT, ora fragment thereof. 5 25. A vector as claimed in claim 17, wherein the DNA segment (s) consists of the following sequence of basest CTA CCG CCA GCA CCT CCT GCT CCT GCA TTT GCT CCT GCT CCT GCA CTT GCA CCG GCT CCT CCA AAT CCT AAT CCT CCA AGT CCT CCG AGT CCT CCG AGT CCA CCG ACT CCA CCG CCG ACT CCT CCA AGT CCA CCA GCT CCT CCG AGT CCA CCA CCG AGT CCT CCG AAT CCA CCT CCG AGT CCA CCT CCG AGT CCA CCG TTT CCT CCG GCT CCA CCT CCG AGT CCA CCA TTT CCT CCG ACT TAT AAA GCA AAG CCA ACT TAT AAA GCA AAG CCA ACT AAT CCT TCA ACG TAT AAA GCA AAG CCA AGT TAT CCT CCA ACT TAT AAA GCA AAA CCA AGT TAT CCT CCA ACT TAT AAA GCA AAA CCA AGT TAT CCT CCA ACG TAT AAA GCA AAG CCA ACT TAT AAA ATA AAG CCA ACT TAT CCT TCA ACG TAT AAA GCA AAG CCA ACT AAT CCT TCA ACG TAT AAA GCA AAG CCA AGT TAT CCT CCA ACT TAT AAA GCA AAA CCA AGT TAT CCT CCA ACT TAT AAA GCA AAA CCA AGT TAT CCT CCA ACG TAT AAA GCA AAG CCA ACT TAT AAA ATA AAG CCA ACT TAT CCT TCA ACG TAT AAA GCA AAG CCA ACT TAT AAA GCA AAG CCA ACT TAT CCT CCA ACT TAT AAA GCA AAA CCA AGT TAT CCT CCA ACA TAT AAA CCA AAG CCA AGT TAT CCT CCA ACT TAT AAA TCC AAG TCA ATA TAT CCC TCT TCA TAC AAA CCT AAG AAA ACT TAT CCC CCC ACA TAT AAA CCT AAA CTA ACC TAT CCT CCA ACA TAT AAA CCA AAG CCA AGT TAT CCA CCA TCT TAT AAA CCT AAG ATT ACT TAT CCC TCA ACT TAT AAA TTG AAG CCA AGT TAT CCT CCA ACA TAC AAA TCT AAA ACA AGT TAC CCT CCT ACA TAT AAC AAA AAG ATC AGC TAT CCA TCA TCA TAT AAA GCT AAG ACA AGT TAT CCC CCA GCA TAT AAA CCA ACA AAC AGA TAT, or a fragment thereof. 26. A vector as claimed in claim 17, wherein the DNA segment (a) comprises the following sequence of bases: TAT AAA GCA AAA CCA AGT TAT CCT CCA ACT TAT AAA GCA AAG CCA ACT TAT AAA GCA AAG CCA ACT AAT CCT TCA ACG TAT AAA GCA AAG CCA AGT TAT CCT CCA ACT TAT AAA GCA AAA CCA AGT TAT CCT CCA ACT TAT AAA GCA AAA CCA AGT TAT CCT CCA ACG TAT AAA GCA AAG CCA ACT TAT AAA GCA AAG CCA ACT TAT CCT TCA ACG TAT AAA GCA AAG CCA ACT TAT AAA GCA AAG CCA ACT TAT CCT CCA ACT TAT AAA GCA AAA CCA AGT TAT CCT CCA ACA TAT AAA CCA AAG CCA AGT TAT CCT CCA ACT TAT AAA TCC AAG TCA ATA TAT CCC TCT TCA TAC AAA CCT AAG AAA ACT TAT CCC CCC ACA TAT, or a fragment thereof. 27. A vector as claimed in claim 17, wherein the DNA segment (a) comprises the following sequence of bases: ACT TCA ACT TAT AAA GCA AAA CCA AGT TAT CCT CCA ACT TAT AAA GCA AAG CCA ACT TAC CCT TCA ACG TAT AAA GCA AAA CCA AGT TAT CCT CCA ACT TAT AAA CCT AAG ATA AGT TAT CCT CCA ACT TAT AAA GCA AAA CCA AGT TAT CCT CCA ACT TAT AAA GCA AAA CCA AGT TAT CCT CCA ACT TAT AAA GCA AAG CCA ACT TAT AAA GCA AAG CCA ACT AAT CCT TCA ACG TAT AAA GCA AAG CCA AGT TAT, or a fragment thereof. 28. A vector as claimed in claim 17, comprising the 0_/P_ promoter-operator operably linked to the DNA segment (a). 29. A vector ss claimed in claim 17, which is the plasmid pGX2383. 30. A vector as claimed in claim 17, comprising the GALl-MT-alpha 1 hybrid promoter. 31. A vector as claimed in claim 17, which is the plasmid YpGX285GAL4. 32. A microorganism, capable of expressing a bio¬ adhesive precursor protein, comprising a microorganism which has been transformed with the expression vector of claim 15, 16, 17, 16, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, or 31. 33. The transformant microorganism E. coli GX3015(pGX2383). 34. The transformant microorganism S. cerevisiae AH22(YpGX285GAL4). 35. A method of producing a bioadhesive protein which comprises: (a) culturing a transformant microorganism that has been transformed with an expression vector comprising: (i) a DNA segment comprising codons for the native amino acid sequence of a bioadhesive precursor protein of a marine animal selected from the group consisting of mussels, bar¬ nacles, and oysters; and -88- ( ii ) a promoter and transcription ini¬ tiation signals , operably linked to said DNA segment, which are capable of effecting microbial expression of the bioadhesive precursor pro¬ tein; ( b) expressing the bioadhesive precursor protein and recovering it from the transformant microorganism; and ( c ) hydroxylating the bioadhesive protein. 36. The method of claim 35 wherein the bioadhe¬ sive precursor protein is from mussel. 37. The method as claimed in claim 36 , wherein the bioadhes ive precursor protein is of the mussel species Mytilus edulis . 38. A method as claimed in claim 37 , wherein the bioadhesive precursor protein contains the amino acid sequence: Lys Leu Ser Ser Tyr Lys Pro He Lys Thr Thr Tyr Asn Ala Lys Thr Asn Tyr Pro Pro Val Tyr Lys Pro Lys Met Thr Tyr Pro Pro Thr Tyr Lys Pro Lys Pro Ser Tyr Pro Pro Thr Tyr Lys Ser Lys Pro Thr Tyr Lys Pro Lys He Thr Tyr Pro Pro Thr Tyr Lys Ala Lys Pro Ser Tyr Pro Pro Thr Tyr Lys Pro Lys Lys Thr Tyr Pro Pre Thr Tyr Lys Pro Lys Leu Thr Tyr Pro Pro Thr Tyr Lys Pro Lys Pro Ser Tyr Pro Pro Thr Tyr Lys Ser Lys Pro Thr Tyr Lys Pro Lys l e Thr Tyr Pre Pro Thr Tyr Lys Ala Lys Pro Ser Tyr Pro Pro Thr Tyr Lys Pro Lys Lys Thr Tyr Pro Pro Thr Tyr Lys Pro Lys Leu Thr Tyr Pro Pro Thr Tyr Lys Pro Lys Ala Ser Tyr Pro Pro Thr Tyr Lys Pro Lys Pro Ser Tyr 89 Pro Pro Ser Tyr Lys Thr Lys Lys Thr Tyr Pro Pro Thr Tyr Lys Pro Lys Leu Thr Tyr Pro Pro Thr Tyr Lys Pro Lys Pro Ser Tyr Pro Pro Ser Tyr Lys Pro Lys Lys Thr Ty Pro Pro Thr Tyr Lys Pro Lys Leu Thr Tyr Pro Pro Thr Tyr Lys Ala Lys Pro Ser Tyr Pro Pro Thr Tyr Lys Ala Lys Pro Ser Tyr Pro Pro Thr Tyr Lys AU Lys Pro Ser Tyr Pro Pro Thr Tyr Lys Ala Lys Pro Str Tyr Pro Ser Thr Tyr Lys Ala Lys Pro Ser Tyr Pro Pro Thr Tyr Lys Ala Lys Pro Thr Tyr Lys Ala Lys Pre Thr Tyr Lys Ala Lys Pro Thr Tyr Pro Str Thr Tyr Lys Ala Lys Pro Thr Tyr Pro Pro Thr Tyr Lys AU Lys Pro Str Tyr Pro Pro Thr Tyr Lys AU Lys Pro Thr Tyr Lys Ala Lys Pro Thr Tyr Lys AU Lys Pro Ser Tyr Pro Pro Thr Tyr Lys AU Lys Pro Ser Tyr Pro Pro Thr Tyr Lys AU Lys Pro Ser Tyr Pro Pro Thr Tyr Lys AU Lys Pro Ser Tyr Pro Pro Thr Tyr Lys AU Lys Pro Str Tyr Pro Pre Thr Tyr Lys Val Lys Pro Thr Tyr Lys Ala Lys Pro Thr Tyr Pro Ser Thr Tyr Lys Ala Lys Pro Ser Tyr Pro Pro Thr Tyr Lys AU Lys Pro Ser Tyr Pro Pro Thr Tyr Lys Ala Lys Pro Ser Tyr Pro Pro Thr Tyr Lys Ala Lys Pro Ser Tyr Pro Pro Thr Tyr Lys Ala Lys Pro Thr Tyr Pro Ser Thr Tyr Lys Ala Lys Pro Ser Tyr Pro Pro Thr Tyr Lys Pro Lys lie Ser Tyr Pro Pro Thr Tyr Lys Ala Lys Pro Ser Tyr Pro Ser Thr Tyr Lys Ala Lys Ser Ser Tyr Pro Pro Thr Tyr Lys Ala Lys Pro Ser Tyr Pro Pro Thr Tyr Lys Ala Lys Pro Thr Tyr Pro Ser Thr Tyr Lys Ala Lys Pro Thr Tyr Lys Ala Lys Pro Ser Tyr Pro Pro Thr Tyr Lys Ala Lys Pro Ser Tyr Pro Pro Thr Tyr Lys Ala Lys Pro Ser Tyr Pro Pro Thr Tyr Lys Ala Lys Pro Thr Tyr Lys Ala Lys Pro Thr Tyr Lys Ala Lys Pro Thr Tyr Pro Ser Thr Tyr Lys Ala Lys Pro Ser Tyr Pro Pro Thr Tyr Lys Ala Lys Pro Ser Tyr Pro Pro Thr Tyr Lys Ala Lys Pro Ser Tyr Pro Pro Thr Tyr Lys Ala Lys Pro Ser Tyr Pro Pro Thr Tyr Lys Ala Lys Pro Ser Tyr Pro Pro Thr Tyr Lys Ala Lys Pro Ser Tyr Pro Pro Thr Tyr Lys Ala Lys Pro Ser Tyr Pro Pro Thr Tyr Lys Ala Lys Pro Thr Tyr Lys Ala Lys Pro Thr Tyr Pro Ser Thr Tyr Lys Ala Lys Pro Ser Tyr Pro Pro Thr Tyr Lys Ala Lys Pro Ser Tyr Pro Pro Thr Tyr Lys Ala Lys Pro Ser Tyr Pro Pro Thr Tyr Lys AU Lys Pro Thr Tyr Lys Ala Lys Pro Thr Tyr Pro Ser Thr Tyr Lys Ala Lys Pro Ser Tyr Pro Pro Thr His Lys Ala Lys Pro Thr Tyr Lys Ala Lys Pro Thr Tyr Pro Ser Thr Tyr Lys Ala Lys Pro Thr Tyr Pro Ser Thr Asp Gly Ala Lys Str, or a fragment thereof. 39. A method as claimed in claim 37, wherein the bioadhesive precursor protein contains the amino acid sequence: Lys Pro Lys Pro Ser Tyr Pro Pro Ser Tyr Lys Pro Lys Thr Thr Tyr Pro Pro Thr Tyr Lys Pro Lys He Ser Tyr Pro Pro Thr Tyr Lys Ala Lys Pro Ser Tyr Pro Ala Thr Tyr Lys Ala Lys Pro Ser Tyr Pro Pro Thr Tyr Lys Ala Lys Pro Ser Tyr Pro Pro Thr Tyr Lys Ala Lys Pro Ser Tyr Pro Pro Thr Tyr Lys Ala Lys Pro Thr Tyr Lys Ala Lys Pro Thr Tyr Pro Pro Thr Tyr Lys Ala Lys Pro Ser Tyr Pro Pro Thr Tyr Lys Pro Lys Pro Ser Tyr Pro Pro Thr Tyr Lys Ser Lys Ser He Tyr Pro Ser Ser Tyr Lys Pro Lys Lys Thr Tyr Pro Pro Thr Tyr Lys Pro Lys Leu Thr Tyr Pro Pro Thr Tyr Lys Pro Lys Pro Ser Tyr Pro Pro Ser Tyr Lys Pro Lys He Thr Tyr Pro Ser Thr Tyr Lys Leu Lys Pro Ser Tyr Pro Pre Thr Tyr Lys Ser Lys Thr Ser Tyr Pro Pro Thr Tyr Asn Lys Lys He Ser Tyr Pro Ser Ser Tyr Lys Ala Lys Thr Ser Tyr Pro Pro Ala Tyr Lys Pro Thr Asn Arg Tyr or a fragment thereof. 91 40. A method as claimed in claim 37, wherein the bioadhesive precursor protein contains the amino acid sequence: Leu .Pro Pro Ala Pro Pro Ala Pro Ala Phe Ala Pro Ala Pro Ala Leu Ala Pro Ala Pro Pro Asn Pro Asn Pro Pro Ser Pro Pro Ser fro Pro Ser Pro Pro Thr Pro Pro Pro Thr Pro Pro tUt Pro Pro Ala Pro Pro Ser Pro Pro Pro Ser Pro Pro Asn Pro Pro Pro Ser Pro Pro Pro Ser Pro Pro Phe Pro Pro Ala Pro Pro Pro Ser Pro Pro Phe Pro Pro Thr Tyr Lys Ala Lys Pro Thr Tyr Lys Ala Lys Pro Thr Asn Pro Ser Thr Tyr Lys Ala Lys Pro Ser Tyr Pro Pro Thr Tyr Lys Ala Lys Pro Ser Tyr Pro Pro Thr Tyr Lys Ala Lys Pro Ser Tyr Pro Pro Thr Tyr Lys Ala Lys Pro Thr Tyr Lys He Lys Pro Thr Tyr Pro Ser Thr Tyr Lys Ala Lys Pro Thr Asn Pro Ser Thr Tyr Lys Ala Lys Pro Ser Tyr Pro Pro Thr Tyr Lys Ala Lys Pro Ser Tyr Pro Pro Thr Tyr Lys Ala Lys Pro Ser Tyr Pro Pro Thr Tyr Lys Ala Lys Pro Thr Tyr Lys lie Lys Pro Thr Tyr Pro Ser Thr Tyr Lys Ala Lys Pro Thr Tyr Lys Ala Lys Pro Thr Tyr Pro Pro Thr Tyr Lys Ala Lys Pro Ser Tyr Pro Pro Thr Tyr Lys Pro Lys Pro Ser Tyr Pro Pro Thr Tyr Lys Ser Lys Ser He Tyr Pro Ser Ser Tyr Lys Pro Lys Lys Thr Tyr Pro Pro Thr Tyr Lys Pro Lys Leu Thr Tyr Pro Pro Thr Tyr Lys Pro Lys Pro Ser Tyr Pro Pro Ser Tyr Lys Pro Lys He Thr Tyr Pro Ser Thr Tyr Lys Leu Lys Pro Ser Tyr Pro Pro Thr Tyr Lys Ser Lys Thr Ser Tyr Pro Pro Thr Tyr Asn Lys Lys He Ser Tyr Pro Ser Ser Tyr Lys Ala Lys Thr Ser Tyr Pro Pro Ala Tyr Lys Pro Thr Asn Arg Tyr or a fragment thereof. 41. A method as claimed in claim 37 wherein the bioadhesive precursor protein contains the amino acid sequence: Tyr Lys Ala Lys Pro Ser Tyr Pro Pro Thr Tyr Lys Ala Lys Pro Thr Tyr Lys Ala Lys Pro Thr Asn Pro Ser Thr Tyr Lys Ala Lys Pro Ser Tyr Pro Pro Thr Tyr Lys Ala 92 Lys Pro Ser Tyr Pro Pro Thr Tyr Lys Ala Lys Pro Ser Tyr Pro Pro Thr Tyr Lys Ala Lys Pro Thr Tyr Lys Ala Lys Pro Thr Tyr Pro Ser Thr Tyr Lys Ala Lys Pro Thr Tyr Lys Ala Lys Pro Thr Tyr Pro Pro Thr Tyr Lys Ala Lys Pro Ser Tyr Pro Pro Thr Tyr Lys Pro Lys Pro Ser Tyr Pro Pro Thr Tyr Lys Ser Lys Ser ∑le Tyr Pro Ser Ser Tyr Lys Pro Lys Lys Thr Tyr Pro Pro Thr Tyr, or a fragment thereof. 42. A method as claimed in claim 37 wherein the bioadhesive precursor protein contains the amino acid sequence: Thr Ser Thr Tyr Lys Ala Lys Pro Ser Tyr Pro Pro Thr » Tyr Lys Ala Lys Pro Thr Tyr Pro Ser Thr Tyr Lys Ala Lys Pro Ser Tyr Pro Pro Thr Tyr Lys Pro Lys He Ser Tyr Pro Pro Thr Tyr Lys Ala Lys Pro Ser Tyr Pro Pro Thr Tyr Lys Ala Lys Pro Ser Tyr Pro Pro Thr Tyr Lys Ala Lys Pro Thr Tyr Lys Ala Lys Pro Thr Asn Pro Ser Thr Tyr Lys Ala Lys Pro Ser Tyr 43. A method as claimed in claim 37, wherein the DNA segment coding for the bioadhesive precursor pro¬ tein comprises the following DNA sequence: MG CTG TCA TCT TAC AM CCT ATT MG ACA ACA TAT MT GCA MG ACA MT TAT CCA CCA 6TT TAT AM CCT MG ATG ACT TAT CCT CCT ACA TAC MA CCA MG CCC AGT TAT CCT CCA ACA TAT AM TCA MG CCC ACA TAC AM CCT MG ATA ACA TAC CCT CCA ACT TAT AAA GCA 7/03048 93 MG CCC AGT TAT CCT CCA ACA TAT AM CCT MG MA ACT TAT CCC CCC ACA TAT MA CCT MA CTA ACC TAT CCT CCT ACA TAC AM CCA MG CCC AGT TAT CCT CCA ACA TAT AM TCA MG CCC ACA TAC AM CCT MG ATA ACA TAC CCT CCA ACA TAT AM GCA MG CCA AGT TAT CCT CCT ACA TAT AM CCT MG MA ACT TAT CCC CCC ACA TAT AM CCT AM CTA ACC TAT CCT CCT ACA TAT AM CCA MG 6CC AGT TAT CCT CCA ACA TAT AM CCA AM CCA AGT TAT CCC CCT TCA TAT MA ACT MG MA ACT TAT CCC CCC ACA TAT AM CCT MA CTA ACC TAT CCT CCT ACA TAT AM CCA AM CCA AGT TAT CCC CCT TCA TAC AM CCT MG MA ACT TAT CCC CCC ACA TAT MA CCT AM CTA ACC TAT CCT CCT ACA TAT AM GCA MG CCA AGT TAT CCT CCA ACA TAT AM GCA AM CCA AGT TAT CCT CCA ACT TAT AM GCA AM CCA AGT TAT CCT CCA ACA TAT MA GCA AM CCA AGT TAT CCT TCA ACT TAT AM GCA AM CCA AGT TAT CCT CCA ACT TAT AM GCA MG CCA ACT TAT AM GCA AAG CCA ACT TAT AM GCA MG CCA ACT TAT CCT TCA ACG TAT AM GCA MG CCA ACT TAT CCT CCA ACT TAT AM GCA MA CCA AGT TAT CCT CtA ACT TAT AM GCA MG CCA ACT TAT AM GCA MG CCA ACT TAT AM GCA MG CCA AGT TAT CCT CCA ACT TAT AM GCA AM CCA AGT TAT CCT CCA ACT TAT AM GCA 94 MG CCA AGT TAT CCT CCA ACG TAT AM GCA MG CCA AGT TAT CCT CCA ACT TAT AM GCA MA CCA AGT TAT CCT CCA ACT TAT AM GTA MG CCA ACA TAT AM GCA AM CCA ACT TAT CCT TCA ACG TAT AM GCA MG CCA AGT TAT CCT CCA ACT TAT AM GCA AM CCA AGT TAT CCT CCA ACT TAT AM GCA AM CCA AGT TAT CCT CCA ACG TAT MA GCA AM CCA AGT TAT CCT CCA ACT TAT AM GCA AM CCA ACT TAT CCT TCA ACG TAT MA GCA AM CCA AGT TAT CCT CCA ACT TAT AM CCT MA ATA AGT TAT CCT CCA ACT TAT AM GCA AM CCA AGT TAT CCT TCA ACT TAT AM GCA MA TCA AGT TAT CCT CCA ACT TAT AM GCA MA CCA AGT TAT CCT CCA ACT TAT MA GCA MG CCA ACT TAT CCT TCA ACG TAT AM GCA MG CCA ACT TAT AM GCA MG CCA AGT TAT CCT CCA ACG ΪAT AM GCA MG CCA AGT TAT CCT CCA ACT TAT AM GCA MA CCC AGT TAT CCT CCA ACT TAT AM GCA MG CCA ACT TAT AM GCA MG CCA ACT TAT AM GCA MG CCA ACT TAT CCT TCA ACG TAT AM GCA MG CCA AGT TAT CCT CCA ACT TAT AM GCA AM CCA AGT TAT CCT CCA ACT TAT AM GCA AM CCA AGT TAT CCT CCA ACT TAT AM GCA AM CCA AGT TAT CCT CCA ACT TAT AM GCA MG CCA AGT TAT CCT CCA ACG TAT AM GCA MG CCA AGT TAT CCT CCA ACT TAT AM GCA AM CCA AGT TAT CCT CCA ACT TAT AM GCA MG CCA ACT TAT AM GCA MG CCA ACT TAT CCT TCA ACG TAT AM GCA MG CCA AGT TAT CCT CCA ACT TAT AM GCA AM tϋA AGT TAT CCT CCA ACT TAT MA GCA MA CCA AGT TAT CCT CCA ACT TAT AM GCA AM CCA ACT TAT AM GCA AAG CCA ACT TAT 95 CCT TCA ACG TAT AM 6CA AM CCA AGT TAT CCT CCA ACT CAT AM GCA AM CCA ACT TAT MA GCA MG CCA ACT TAT CCT TCA ACT TAT AM GCA AM CCA ACT TAT CCT TCA ACG GAT 6GA GCA AM TCA. or a fragment thereof. 44. A method as claimed in claim 37, wherein the DNA segment coding for the bioadhesive precursor pro¬ tein comprises the following DNA sequence: AAA CCA AAA CCA AGT TAT CCA CCA TCT TAT AAA CCT AAA ACA ACT TAT CCT CCA ACT TAT AAA CCT AAG ATA AGT TAT CCT CCA ACT TAT AAA GCA AAA CCA AGT TAT CCA GCA ACT TAT AAA GCA AAA CCA AGT TAT CCT CCA ACT TAT AAA GCA AAA CCA AGT TAT CCT CCA ACT TAT AAA GCA AAA CCA AGT TAT CCT CCA ACT TAT AAA GCA AAG CCA ACT TAT AAA GCA AAG CCA ACT TAT CCT CCA ACT TAT.AAA GCA AAA CCA AGT TAT CCT CCA ACA TAT AAA CCA AAG CC AGT TAT CCT CCA ACT TAT AA\ TCC AAG TCA ATA TAT CCC TCT TCA TAC AAA CCT AAG AAA ACT TAT CCC CCC ACA TAT AAA CCT AAA CTA ACC TAT CCT CCA ACA TAT AAA CCA AAG CCA AGT TAT CCA CCA TCT TAT AAA CCT AAG ATT ACT TAT CCC TCA ACT TAT AAA TTG AAG CCA AGT TAT CCT CCA ACA TAC AAA TCT AAA ACA AGT TAC CCT CCT ACA TAT AAC AAA AAG ATC AGC TAT CCA TCA TCA TAT AAA GCT AAG ACA AGT TAT CCC CCA GCA TAT AAA CCA ACA AAC ACA TAT, or a fragment thereof. 45. A method ss claimed in claim 37, wherein the DNA segment coding for the bioadhesive precursor pro¬ tein comprises the following DNA sequences: CTA CCG CCA GCA CCT CCT GCT CCT GCA TTT GCT CCT GCT CCT GCA CTT GCA CCG GCT CCT CCA AAT CCT AAT CCT CCA AGT CCT CCG AGT CCT CCG AGT CCA CCG ACT CCA CCG CCG ACT CCT CCA AGT CCA CCA GCT CCT CCG AGT CCA CCA CCG AGT CCT CCG AAT CCA CCT CCG AGT CCA CCT CCG AGT CCA CCG TTT CCT CCG GCT CCA CCT CCG AGT CCA CCA TTT CCT CCG ACT TAT AAA GCA AAG CCA ACT TAT AAA GCA AAG CCA ACT AAT CCT TCA ACG TAT AAA GCA AAG CCA AGT TAT CCT CCA ACT TAT AAA GCA AAA CCA AGT TAT CCT CCA ACT TAT AAA GCA AAA CCA AGT TAT CCT CCA ACG TAT AAA GCA AAG CCA ACT TAT AAA ATA AAG CCA ACT TAT CCT TCA ACG. TAT AAA GCA AAG CCA ACT AAT CCT TCA ACG TAT AAA GCA AAG CCA AGT TAT CCT CCA ACT TAT AAA GCA AAA CCA AGT TAT CCT CCA ACT TAT AAA GCA AAA CCA AGT TAT CCT CCA ACG TAT AAA GCA AAG CCA ACT TAT AAA ATA AAG CCA ACT TAT CCT TCA ACG TAT AAA GCA AAG CCA ACT TAT AAA GCA AAG CCA ACT T T CCT CCA ACT TAT AAA GCA AAA CCA AGT TAT CCT CCA ACA TAT AAA CCA AAG CCA AGT TAT CCT CCA ACT TAT AAA TCC AAG TCA ATA TAT CCC TCT TCA TAC AAA CCT AAG AAA ACT TAT CCC CCC ACA TAT AAA CCT AAA CTA ACC TAT CCT CCA ACA TAT AAA CCA AAG CCA AGT TAT CCA CCA TCT TAT AAA CCT AAG ATT ACT TAT CCC TCA ACT TAT AAA TTG AAG CCA AGT TAT CCT CCA ACA TAC AAA TCT AAA ACA AGT TAC CCT CCT ACA TAT AAC AAA AAG ATC AGC TAT CCA TCA TCA TAT AAA GCT AAG ACA AGT TAT CCC CCA GCA TAT AAA CCA ACA AAC AGA TAT, or a fragment thereof. 46. A method as claimed in claim 37, wherein the DNA segment coding for the bioadhesive precursor pro¬ tein comprises the following DNA sequence: TAT AAA GCA AAA CCA AGT TAT CCT CCA ACT TAT AAA GCA AAG CCA ACT TAT AAA GCA AAG CCA ACT AAT CCT TCA ACG TAT AAA GCA AAG CCA AGT TAT CCT CCA ACT TAT AAA GCA AAA CCA AGT TAT CCT CCA ACT TAT AAA GCA AAA CCA AGT TAT CCT CCA ACG TAT AAA GCA AAG CCA ACT TAT AAA GCA AAG CCA ACT TAT CCT TCA ACG TAT AAA GCA AAG CCA ACT TAT AAA GCA AAG CCA ACT TAT CCT CCA ACT TAT AAA GCA AAA CCA AGT TAT CCT CCA ACA TAT AAA CCA AAG CCA AGT TAT CCT CCA ACT TAT AAA TCC AAG TCA ATA TAT CCC TCT TCA TAC AAA CCT AAG AAA ACT TAT CCC CCC ACA TAT, or a fragment thereof. 47. A method as claimed in claim 37, wherein the DNA segment coding for the bioadhesive precursor pro¬ tein comprises the following DNA sequence: ACT TCA ACT TAT AAA GCA AAA CCA AGT TAT CCT CCA ACT TAT AAA GCA AAG CCA ACT TAC CCT TCA ACG TAT AAA GCA AAA CCA AGT TAT CCT CCA ACT TAT AAA CCT AAG ATA AGT TAT CCT CCA ACT TAT AAA GCA AAA CCA AGT TAT CCT CCA ACT TAT AAA GCA AAA CCA AGT TAT CCT CCA ACT TAT AAA GCA AAG CCA ACT TAT AAA GCA AAG CCA ACT AAT CCT TCA ACG TAT AAA GCA AAG CCA AGT TAT, or a fragment thereof. 48. A method as claimed in claim 37, wherein the expression vector is the plasmid pGX2383. 49. A method as claimed in claim 37, wherein the expression vector is the plasmid YpGX285GAL4. 50. A method as claimed in claim 37, wherein the transformant microorganism is a gram-negative microorganism. 51. A method as claimed in claim S0» wherein the transformant microorganism is E. coli 6X3015(pGX2383 ) . 52. A method as claimed in claim 37, wherein the transformant microorganism is a yeast. 53. A method according to claim 52, wherein the transformant microorganism is Saccharomyces. 54. A method according to claim 53, wherein the transformant microorganism is S. cerevisiae. 55. A method as claimed in claim 54, wherein the transformant microorganism is S. cerevisiae AH22CYpGX285GAL4). 56. A method as claimed in claim 37, wherein the bioadhesive precursor protein is hydroxylated enzyma- tically by treatment with mushroom tyrosinase or Streptomyces antiblotlcus tyrosinase. 57. A method as claimed in claim 56, wherein the bioadhesive precursor protein is hydroxylated by treatment with mushroom tyrosinase. 58. A method of bonding two surfaces which com¬ prises hydroxylating a microbially produced bioadhe¬ sive precursor protein comprising the native amino acid sequence of a bioadhesive precursor protein of a marine animal selected from the group consisting of mussels, barnacles, and oysters or a fragment thereof; applying the hydroxylated, microbially produced bioad¬ hesive precursor protein to one or both surfacesι and contacting the surfaces. 59. The method of claim 58, wherein the bioadhe¬ sive precursor protein is of mussel. 60. The method of claim 59, wherein the bioadhe¬ sive precursor protein is of the mussel species Mytil¬ us edulis. €1. A method as claimed in claim 60, wherein the bioadhesive precursor protein contains the amino acid sequence: Lys Leu Ser Ser Tyr Lys Pro lie Lys Thr Thr Tyr Asn AU Lys Thr Asn Tyr Pro Pro Val Tyr Lys Pro Lys Met Thr Tyr Pro Pro Thr Tyr Lys Pro Lys Pro Ser Tyr Pro Pro Thr Tyr Lys Ser Lys Pro Thr Tyr Lys Pro Lys lie Thr Tyr Pro Pro Thr Tyr Lys Ala Lys Pro Str Tyr Pro Pro Thr Tyr Lys Pro Lys Lys Thr Tyr Pro Pro Thr Tyr Lys Pro Lys Leu Thr Tyr Pro Pro Thr Tyr Lys Pro Lys Pro Ser Tyr Pro Pro Thr Tyr Lys Ser Lys Pro Thr Tyr Lys Pro Lys lit Thr Tyr Pro Pro Thr Tyr Lys AU Lys Pro Ser Tyr Pro Pro Thr Tyr Lys Pro Lys Lys Thr Tyr Pro Pro Thr Tyr Lys Pro Lys Leu Thr Tyr Pro Pro Thr Tyr Lys Pro Lys AU Str Tyr Pro Pro Thr Tyr Lys Pro Lys Pro Ser Tyr Pro Pro Ser Tyr Lys Thr Lys Lys Thr Tyr Pro Pro Thr Tyr Lys Pro Lys Leu Thr Tyr Pro Pro Thr Tyr Lys Pro Lys Pro Str Tyr Pro Pro Ser Tyr Lys Pro Lys Lys Thr Tyr Pro Pro Thr Tyr Lys Pro Lys Ltu Thr Tyr Pro Pro Thr Tyr Lys A Lys Pro Ser Tyr Pro Pro Thr Tyr Lys AU Lys Pro Ser Tyr Pro Pro Thr Tyr Lys AU Lys Pro Ser Tyr Pro Pro Thr Tyr Lys AU Lys Pro Str Tyr Pro Str Thr Tyr Lys AU Lys Pre Ser Tyr Pre Pro Thr Tyr Lys Ala Lys Pro Thr Tyr Lys AU Lys Pro Thr Tyr Lys AU Lys Pro Thr Tyr Pro Str Thr Tyr Lys Ala Lys Pro Thr Tyr Pro Pre Thr Tyr Lys Ala Lys Pre Str Tyr Pro Pro Thr Tyr Lys AU Lys Pro Thr Tyr Lys Ala Lys Pro Thr Tyr Lys AU Lys Pro Ser Tyr Pro Pro Thr Tyr Lys AU Lys Pro Str Tyr Pro Pro Thr Tyr Lys A Lys Pro Ser Tyr Pro Pro Thr Tyr Lys AU Lys Pro Str Tyr Pro Pro Thr Tyr Lys AU Lys Pro Ser Tyr Pre Pro Thr Tyr Lys Val Lys Pro Thr Tyr Lys AU Lys Pro Thr Tyr Pro Ser Thr Tyr Lys AU Lys Pro Ser Tyr Pro Pro Thr Tyr Lys AU Lys Pro Ser Tyr Pre Pro Thr Tyr Lys Ala Lys Pre Ser Tyr Pro Pro Thr Tyr Lys Ala Lys Pro Ser Tyr Pre Pre Thr Tyr Lys Ala Lys Pro Thr Tyr Pro Ser Thr Tyr Lys AU Lys Pre Ser Tyr Pro Pre Thr Tyr Lys Pre Lys ϊ e Ser Tyr Pro Pro Thr Tyr Lys AU Lys Pre Ser Tyr Pro Ser Thr Tyr Lys AU Lys Ser Ser Tyr Pro Pro Thr Tyr Lys AU Lys Pro Ser Tyr Pro Pro Thr Tyr Lys Ala Lys Pro Thr Tyr Pro Ser Thr Tyr Lys Ala Lys Pro Thr Tyr Lys A Lys Pro Ser Tyr Pre Pro Thr Tyr Lys Ala Lys Pro Ser Tyr Pro Pro Thr Tyr Lys Ala Lys Pro Ser Tyr Pro Pro Thr Tyr Lys Ala Lys Pro Thr Tyr Lys Ala Lys Pro Thr Tyr Lys Al a Lys Pro Thr Tyr Pro Ser Thr Tyr Lys Al a Lys Pro Ser Tyr Pro Pro Thr Tyr Lys Ala Lys Pro Ser Tyr Pro Pro Thr Tyr Lys Ala Lys Pre Ser Tyr Pro Pro Thr Tyr Lys Ala Lys Pro Ser Tyr Pro Pro Thr Tyr Lys Al a Lys Pro Ser Tyr Pro Pro Thr Tyr Lys AU Lys Pro Ser Tyr Pro Pro Thr Tyr Lys Ala Lys Pro Ser Tyr Pro Pre Thr Tyr Lys AU Lys Pro Thr Tyr Lys AU Lys Pro Thr Tyr Pro Ser Thr Tyr Lys Ala Lys Pro Ser Tyr Pro Pro Thr Tyr Lys AU Lys Pro Ser Tyr Pro Pro Thr Tyr Lys Al a Lys Pro Ser Tyr Pro Pro Thr Tyr Lys Ala Lys Pro Thr Tyr Lys Ala Lys Pro Thr Tyr Pro Ser Thr Tyr Lys Ala Lys Pre Ser Tyr Pro Pro Thr His Lys Ala Lys Pro Thr Tyr Lys AU Lys Pro Thr Tyr Pro Ser Thr Tyr Lys Al a Lys Pro Thr Tyr Pro Ser Thr Asp Gly Ala Lys Ser, or a fragment thereof. -101- 62. A method as claimed in claim 60, wherein the bioadhesive precursor protein has the amino acid se¬ quence: Lys Pro Lys Pro Ser Tyr Pro Pro Ser Tyr Lys Pro Lys Thr Thr Tyr Pro Pro Thr Tyr Lys Pro Lys He Ser Tyr Pro Pro Thr Tyr Lys Ala Lys Pro Ser Tyr Pro Ala Thr Tyr Lys Ala Lys Pro Ser Tyr Pro Pro Thr Tyr Lys Ala Lys Pro Ser Tyr Pro Pro Thr Tyr Lys Ala Lys Pro Ser Tyr Pro Pro Thr Tyr Lys Ala Lys Pro Thr Tyr Lys Ala Lys Pro Thr Tyr Pro Pro Thr Tyr Lys Ala Lys Pro Ser Tyr Pro Pro Thr Tyr Lys Pre Lys Pro Ser Tyr Pro Pro Thr Tyr Lys Ser Lys Ser He Tyr Pro Ser Ser Tyr Lys Pro Lys Lys Thr Tyr Pro Pro Thr Tyr Lys Pro Lys Leu Thr Tyr Pro Pro Thr Tyr Lys Pro Lys Pro Bet Tyr Pro Pro Ser Tyr Lys Pro Lys He Thr Tyr Pro Ser Thr Tyr Lys Leu Lys Pro Ser Tyr Pro Pro Thr Tyr Lys Ser Lys Thr Ser Tyr Pro Pro Thr Tyr Asn Lys Lys He Ser Tyr Pro Ser Ser Tyr Lys Ala Lys Thr Ser Tyr Pro Pro Ala Tyr Lys Pro Thr Asn Arg Tyr or a fragment thereof. . . 102 S3. A method ss claimed in claim 60, wherein the bioadhesive precursor protein has the amino acid se¬ quences .Leu Pro Pro Ala Pro Pro Ala Pro Ala Phe Aia Pro Ala Pro Ala Leu Ala Pro Ala Pro Pre Asn Pro Asn Pro Pro Ser Pro Pro Ser Pro Pro Ser Pro Pro Thr Pro Pro Pro Thr Pro Pro Ser Pro Pro Ala Pro Pre Ser Pro Pro Pro Ser Pro Pro Asn Pro Pro Pro %e t Pro Pro Pro Ser Pro Pro Phe Pro Pro Ala Pro Pro Pro Ser Pro Pro Phe Pro Pro Thr Tyr Lys Ala Lys Pro Thr Tyr Lys Ala Lys Pro Thr Asn Pro Ser Thr Tyr Lys Ala Lys Pro Ser Tyr Pro Pro Thr Tyr Lys Ala Lys Pro Ser Tyr Pro Pro Thr Tyr Lys Ala Lys Pro Ser Tyr Pro Pro Thr Tyr Lys Ala Lys Pro Thr Tyr Lys He Lys Pro Thr Tyr Pro Ser Thr Tyr Lys Ala Lys Pro Thr Asn Pro Ser Thr Tyr Lys Ala Lys Pro Ser Tyr Pre Pro Thr Tyr Lys Ala Lys Pro Ser Tyr Pro Pro Thr Tyr Lys Ala Lys Pro Ser Tyr Pro Pro Thr Tyr Lys Ala Lys Pro Thr Tyr Lys He Lys Pro Thr Tyr Pro Ser Thr Tyr Lys Ala Lys Pro Thr Tyr Lys Ala Lys Pro-Thr Tyr Pro Pro Thr Tyr Lys Ala Lys Pro Ser Tyr Pro Pro Thr Tyr Lys Pro Lys Pro Ser Tyr Pro Pro Thr Tyr Lys Ser Lys Ser He Tyr Pro Ser Ser Tyr Lys Pro Lys Lys Thr Tyr Pro Pro Thr Tyr Lys Pro Lys Leu Thr Tyr Pro Pro Thr Tyr Lys Pre Lys Pro Ser Tyr Pro Pro Ser Tyr Lys Pro Lys He Thr Tyr Pro Ser Thr Tyr Lys Leu Lys Pro Ser Tyr Pro Pro Thr Tyr Lys Ser Lys Thr Ser Tyr Pro Pro Thr Tyr Asn Lys Lys He Ser Tyr Pro Ser Ser Tyr Lys Ala Lys Thr Ser Tyr Pro Pro Ala Tyr Lys Pro Thr Asn Arg Tyr or a fragment thereof. -103- 64. A method ss claimed in claim 60 wherein the bioadhesive precursor protein has the amino acid se¬ quence: Tyr Lys Ala Lys Pro Ser Tyr Pro Pro Thr Tyr Lys Ala Lys Pro Thr Tyr Lys Ala Lys Pro Thr Asn Pro Ser Thr Tyr Lys Ala Lys Pro Ser Tyr Pro Pro Thr Tyr Lys Ala Lys Pro Ser Tyr Pro Pro Thr Tyr Lys Ala Lys Pro Ser Tyr Pro Pro Thr Tyr Lys Ala Lys Pro Thr Tyr Lys Ala Lys Pro Thr Tyr Pro Ser Thr Tyr Lys Ala Lys Pro Thr Tyr Lys Ala Lys Pro Thr Tyr Pro Pro Thr Tyr Lys Ala Lys Pro Ser Tyr Pro Pro Thr Tyr Lys Pro Lys Pro Ser Tyr Pro Pro Thr Tyr Lys Ser Lys Ser He Tyr Pro Ser Ser Tyr Lys Pro Lys Lys Thr Tyr Pro Pro Thr Tyr, or a fragment thereof. 65. A method as claimed in claim 60 wherein the bioadhesive precursor protein has the amino acid se¬ quence: Thr Ser Thr Tyr Lys Ala Lys Pro Ser Tyr Pro Pro Thr Tyr Lys Ala Lys Pro Thr Tyr Pro Ser Thr Tyr Lys Ala Lys Pro Sex Tyr Pro Pro Thr Tyr Lys Pro Lys He Ser Tyr Pro Pro Thr Tyr Lys Ala Lys Pro Ser Tyr Pro Pro Thr Tyr Lys Ala Lys Pro Ser Tyr Pro Pro Thr Tyr Lys Ala Lys Pro Thr Tyr Lys Ala Lys Pro Thr Asn Pro Ser Thr Tyr Lys Ala Lys Pro Ser Tyr, or a fragment thereof. 66. A method as claimed in claim 58, wherein the bioadhesive precursor protein is hydroxylated, applied to one or both surfaces as a primer coating and an¬ other adhesive is then applied over the primer coat¬ ing. 67. A method as claimed in claim 66, wherein the adhesive that is applied over the primer coating is an epoxy adhesive. 68. A method as claimed in claim 58, wherein the bioadhesive precursor protein is hydroxylated, dis¬ solved in a solvent in admixture with a carbohydrate adhesive and applied to one or both surfaces to be bonded. 69. A method as claimed in claim 68, wherein the carbohydrate adhesive is chitosan. 70. A method of producing a bioadhesive precursor protein which comprises microbially expressing a fu¬ sion protein comprising the native amino acid sequence of a bioadhesive precursor protein of a marine animal selected from the group consisting of mussels, barnacles, and oysters, or a fragment thereof, having an additional amino acid sequence fused to one or both ends thereof, said additional amino acid sequence(s) having a methionine residue within about 10 residues from the bioadhesive precursor protein sequence; and treating the fusion protein with cyanogen bromide to remove all or a portion of the additional amino acid sequence(s) . -105- 71. An adhesive composition comprising: (a) a bioadhesive protein produced by hy- droxylating at least about 10% of the tyrosine residues of a protein having the native amino acid sequence of a bio¬ adhesive precursor protein of a marine animal selected from the group consist¬ ing of mussels, barnacles, and oysters, or a fragment thereof; (b) a carbohydrate adhesive; and (c) a solvent. 72. An adhesive composition as claimed in claim 71, wherein the carbohydrate is chitosan. 73. An adhesive composition as claimed in claim 71, which contains from about 2% to 30% by weight of the adhesive protein and from about 1% to 7% by weight of chitosan. 74. An adhesive composition comprising a solution having 10% to 70% ' solids content, said solids compris¬ ing from about 1% to 50% by weight of a bioadhesive protein produced by hydroxylating at least about 10% of the tyrosine residues of a recombinant protein hav¬ ing the native amino acid sequence of a bioadhesive precursor protein of a mussel of marine animal selected from the group consisting of mussels, barnacles, and oysters, or a fragment thereof; and from about 50% to 99% by weight collagen. -106- 75. An adhesive product comprising a sheet of collagen having on its surfaces an adhesive recombin¬ ant protein produced by hydroxylating at least about 10% of the tyrosine residues of a protein having the native amino acid sequence of a bioadhesive precursor protein of a marine animal selected from the group consisting of mussels, barnacles, and oysters or a fragment thereof.

Inventors: ANDERSON DAVID M, FILPULA DAVID, MAUGH KATHY J, MCCANDLISS RUSS, STRAUSBERG ROBERT, STRAUSBERG SUSAN L, WEI TENA, ANDERSON, DAVID M., FILPULA, DAVID, MAUGH, KATHY J., MCCANDLISS, RUSS, STRAUSBERG, ROBERT, STRAUSBERG, SUSAN L., WEI, TENA
Applicants: GENEX CORP, GENEX CORPORATION
Application year: 1987
Main classification: C12P21
IPC codes: C07K14, C07K19, C09J189, C12N1, C12N9, C12N15, C12P21, C12R1