The Structure and Assembly of Secreted Mucins (1999)

porcine submaxillary mucin bovine submaxillary mucin canine tracheobronchial mucin frog integumentary mucin mouse submandibular mucin porcine gastric mucin rat submandibular mucin human von Willebrand factor Mucins are major glycoprotein components of the mucous that coats the surfaces of cells lining the respiratory, digestive, and urogenital tracts, and in some amphibia, the skin. They function to protect epithelial cells from infection, dehydration, and physical or chemical injury, as well as to aid the passage of materials through a tract. Individual organisms make several structurally different mucins, and a given mucin may be found in more than one organ (see Supplemental Material). Members of the mucin family can differ considerably in size. Some are small, containing a few hundred amino acid residues, whereas others contain several thousands of residues and are among the largest known proteins. Irrespective of size, all mucin polypeptide chains have domains rich in threonine and/or serine whose hydroxyl groups are in O-glycosidic linkage with oligosaccharides. Moreover, these domains are composed of tandemly repeated sequences that vary in number, length, and amino acid sequence from one mucin to another (1Gendler S.J. Spicer A.P. Annu. Rev. Physiol. 1995; 37: 607-634Crossref Scopus (854) Google Scholar). The carbohydrate content of a mucin may account for up to 90% of its weight. There are two types of mucins, membrane-bound and secreted. Of the human mucins, two are membrane-bound (MUC1 and MUC4) (2Gendler S.J. Lancaster C.A. Taylor-Papadimitriu J. Duhig T. Peat N. Burchell J. Pemberton L. Lalani E.N. Wilson D. J. Biol. Chem. 1990; 265: 15286-15293Abstract Full Text PDF PubMed Google Scholar, 3Moniaux N. Nollet S. Porchet N. Degand P. Laine A. Aubert J.-P. Biochem. J. 1999; 338: 325-333Crossref PubMed Scopus (219) Google Scholar) and four are secreted (MUC2, MUC5AC, MUC5B, and MUC7) (4Gum J.R. Hicks J.W. Toribara N.W. Siddiki B. Kim Y.S. J. Biol. Chem. 1994; 269: 2440-2446Abstract Full Text PDF PubMed Google Scholar, 5Li D. Gallup M. Fan N. Szymkowski D.E. Basbaum C.B. J. Biol. Chem. 1998; 273: 6812-6820Abstract Full Text Full Text PDF PubMed Scopus (156) Google Scholar, 6Desseyn J.L. Aubert J.P. van Seuningen I. Porchet N. Laine A. J. Biol. Chem. 1997; 272: 16873-16883Abstract Full Text Full Text PDF PubMed Scopus (93) Google Scholar, 7Bobek L.A. Tsa H. Biesbrock A.R. Levine M.J. J. Biol. Chem. 1993; 268: 20563-20569Abstract Full Text PDF PubMed Google Scholar). The three other mucins (MUC3, MUC6, and MUC8) (8Gum J.R. Ho J.J.L. Pratt W.S. Hicks J.W. Hill A.S. Vinall L.E. Roberton A.M. Swallow D.M. Kim Y.S. J. Biol. Chem. 1992; 267: 26678-26686Abstract Full Text PDF Google Scholar, 9Toribara N.W. Ho S.B. Gum E. Gum J.R. Lau P. Kim Y.S. J. Biol. Chem. 1997; 272: 16398-16403Abstract Full Text Full Text PDF PubMed Scopus (87) Google Scholar, 10Shankar V. Pichan P. Eddy Jr., R.L. Tonk V. Nowak N. Sait S.N. Shows T.B. Schultz R.E. Gotway G. Elkins R.C. Gilmore M.S. Sachdev G.P. Am. J. Respir. Cell Mol. Biol. 1997; 16: 232-241Crossref PubMed Scopus (128) Google Scholar, 11Pigny P. Guyonnet-Duperat V. Hill A.S. Pratt W.S. Galiegue-Zouitina S. Vinall L. Collyn D'Hooge M. Laine A. Van-Seuningen I. Degand P. Gum J.R. Kim Y.S. Swallow D.M. Aubert J.-P. Porchet N. Genomics. 1996; 38: 340-352Crossref PubMed Scopus (187) Google Scholar) cannot be classified. Each human mucin has a counterpart in other animals. Thus, porcine submaxillary mucin (PSM)1 (12Eckhardt A.E. Timpte C.S. DeLuca A.W. Hill R.L. J. Biol. Chem. 1997; 272: 33204-33210Abstract Full Text Full Text PDF PubMed Scopus (48) Google Scholar), one of the most thoroughly characterized mucins, has a tissue distribution and structure similar to MUC5B. An increasing number of proteins that are not mucins also contain highly O-glycosylated domains called “mucin-like domains.” The functions of mucins are dependent on their ability to form viscous solutions or gels. Although the highly glycosylated domains of mucins are devoid of secondary structures, they are long extended structures that are much less flexible than unglycosylated random coils. The oligosaccharides contribute to this stiffness in two ways, by limiting the rotation around peptide bonds and by charge repulsion among the neighboring, negatively charged oligosaccharide groups (13Jentoft N. Trends Biochem. Sci. 1991; 15: 291-294Abstract Full Text PDF Scopus (621) Google Scholar). Such long, extended molecules have a much greater solution volume than native or denatured proteins with little or no carbohydrate and endow aqueous mucin solutions with a high viscosity. Mucins protect against infection by microorganisms that bind cell surface carbohydrates, and mucin genes appear to be up-regulated by substances derived from bacteria, e.g. lipopolysaccharides (14Dohrman A. Miyata S. Gallup M. Li J.D. Chapelin C. Coste A. Escudier E. Nadel J. Basbaum C. Biochim. Biophys. Acta. 1998; 1406: 251-259Crossref PubMed Scopus (183) Google Scholar). This review will summarize what is known about the polypeptide structures of the secreted mucins and how some, in particular PSM, are assembled via interchain disulfide bonds into molecules with molecular weights in the millions. We will not consider membrane-bound mucins, which were the subject of earlier reviews (1Gendler S.J. Spicer A.P. Annu. Rev. Physiol. 1995; 37: 607-634Crossref Scopus (854) Google Scholar, 15Hilkens J. Marjolyn J.L. Ligtenberg H.L.V. Litvinov S.V. Trends Biochem. Sci. 1992; 17: 359-363Abstract Full Text PDF PubMed Scopus (434) Google Scholar, 16McNeer R.R. Huang D. Fregien N.L. Carraway K.L. Biochem. J. 1998; 330: 737-744Crossref PubMed Scopus (47) Google Scholar). Complete amino acid sequences have been described for frog (Xenopus) integumentary mucins FIM-A.1 (17Hoffmann W. J. Biol. Chem. 1988; 263: 7686-7690Abstract Full Text PDF PubMed Google Scholar) and FIM-B.1 (18Joba W. Hoffmann W. J. Biol. Chem. 1997; 272: 1805-1810Abstract Full Text Full Text PDF PubMed Scopus (21) Google Scholar), PSM (12Eckhardt A.E. Timpte C.S. DeLuca A.W. Hill R.L. J. Biol. Chem. 1997; 272: 33204-33210Abstract Full Text Full Text PDF PubMed Scopus (48) Google Scholar), RSM (19Albone E.F. Hagen F.K. VanWuyckhuyse B.C. Tabak L.A. J. Biol. Chem. 1994; 269: 16845-16852Abstract Full Text PDF PubMed Google Scholar), MSM (20Denny P.C. Mirels L. Denny P.A. Glycobiology. 1996; 6: 43-50Crossref PubMed Scopus (28) Google Scholar), MUC2 (4Gum J.R. Hicks J.W. Toribara N.W. Siddiki B. Kim Y.S. J. Biol. Chem. 1994; 269: 2440-2446Abstract Full Text PDF PubMed Google Scholar), MUC5B (6Desseyn J.L. Aubert J.P. van Seuningen I. Porchet N. Laine A. J. Biol. Chem. 1997; 272: 16873-16883Abstract Full Text Full Text PDF PubMed Scopus (93) Google Scholar), and MUC7 (7Bobek L.A. Tsa H. Biesbrock A.R. Levine M.J. J. Biol. Chem. 1993; 268: 20563-20569Abstract Full Text PDF PubMed Google Scholar) and almost complete sequences for MUC5AC (5Li D. Gallup M. Fan N. Szymkowski D.E. Basbaum C.B. J. Biol. Chem. 1998; 273: 6812-6820Abstract Full Text Full Text PDF PubMed Scopus (156) Google Scholar) and rat Muc2 (21Ohmori H. Dohrman A.F. Gallup M. Tsuda T. Kai H. Gum J.R. Kim Y.S. Basbaum C.B. J. Biol. Chem. 1994; 269: 17833-17840Abstract Full Text PDF PubMed Google Scholar). The different domains of mucins are shown in Fig.1. Many of the domains show sequence identities and possibly similar functions in different mucins. These mucins vary greatly in size, from as few as 322 residues to 13,288 residues. The sequences of mucin polypeptides were deduced almost completely by recombinant DNA methods, and the physical-chemical properties of some mucins have not been determined. Nevertheless, it is well established that the oligosaccharides in many secreted mucins, e.g. PSM (22Gerken T.A. Owens C.L. Pasumarthy M. J. Biol. Chem. 1998; 273: 26580-26588Abstract Full Text Full Text PDF PubMed Scopus (40) Google Scholar), show structural microheterogeneity, with GalNAcα-O-Ser/Thr as the sugar-protein linkage upon which other sugars are added. Most mucins have negatively charged sugars, either sialic acid or O-sulfosaccharides. The number, length, and amino acid sequence of the repeats vary among different mucins, as shown in the Supplemental Material. The tandem repeat domains are flanked on either side by other types of domains (Fig. 1). All of the serine and threonine residues in the repeat domain of PSM have O-linked oligosaccharides (23Gerken T.A. Owens C.L. Pasumarthy M. J. Biol. Chem. 1997; 272: 9709-9719Abstract Full Text Full Text PDF PubMed Scopus (76) Google Scholar), but this is not known for other mucins. The repeats in some mucins have identical sequences, whereas in others the repeat sequence is degenerate. The lack of secondary structures in the repeat domains and their flanking domains suggests that these domains serve as a scaffold forO-linked oligosaccharides (24Eckhardt A.E. Timpte C.S. Abernethy J.L. Toumadje A. Johnson Jr., W.C. Hill R.L. J. Biol. Chem. 1987; 262: 11339-11344Abstract Full Text PDF PubMed Google Scholar), whose properties determine in large part the properties of a mucin. Light scattering and electron microscopy suggest that these glycosylated domains are semi-rigid, extended structures (13Jentoft N. Trends Biochem. Sci. 1991; 15: 291-294Abstract Full Text PDF Scopus (621) Google Scholar, 25Marianne T. Perini J.-M. Lafitte J.-J. Houdret N. Pruvot F.-R. Lamblin G. Slayter H.S. Roussel P. Biochem. J. 1987; 248: 189-195Crossref PubMed Scopus (31) Google Scholar). The tandem repeat domains in many mucins, e.g. PSM (12Eckhardt A.E. Timpte C.S. DeLuca A.W. Hill R.L. J. Biol. Chem. 1997; 272: 33204-33210Abstract Full Text Full Text PDF PubMed Scopus (48) Google Scholar) and MUC5B (6Desseyn J.L. Aubert J.P. van Seuningen I. Porchet N. Laine A. J. Biol. Chem. 1997; 272: 16873-16883Abstract Full Text Full Text PDF PubMed Scopus (93) Google Scholar), are encoded by a single large exon, although the remainder of the mucin is encoded by short exons separated by long introns. Many mucins show length polymorphism as the result of multiple alleles that encode different numbers of tandem repeats (26Vinall L.E. Hill A.S. Pigny P. Pratt W.S. Toribara N. Gum J.R. Kim Y.S. Porchet N. Aubert J.-P. Swallow D.M. Hum. Genet. 1998; 102: 357-366Crossref PubMed Scopus (70) Google Scholar). Thus, PSM is encoded by at least three alleles with 99, 110, and 135 repeats, respectively (12Eckhardt A.E. Timpte C.S. DeLuca A.W. Hill R.L. J. Biol. Chem. 1997; 272: 33204-33210Abstract Full Text Full Text PDF PubMed Scopus (48) Google Scholar). Several disulfide-rich domains are found in secreted mucins except RSM (19Albone E.F. Hagen F.K. VanWuyckhuyse B.C. Tabak L.A. J. Biol. Chem. 1994; 269: 16845-16852Abstract Full Text PDF PubMed Google Scholar), MSM (20Denny P.C. Mirels L. Denny P.A. Glycobiology. 1996; 6: 43-50Crossref PubMed Scopus (28) Google Scholar), and MUC7 (7Bobek L.A. Tsa H. Biesbrock A.R. Levine M.J. J. Biol. Chem. 1993; 268: 20563-20569Abstract Full Text PDF PubMed Google Scholar) and are often at either end of the polypeptide (Fig. 1). The disulfide-rich D-domain in mucins first found in VWF (27Sadler J.E. Annu. Rev. Biochem. 1998; 67: 395-424Crossref PubMed Scopus (1109) Google Scholar) is now recognized in many other proteins (28Kotani E. Yamakawa M. Iwamoto S. Tashiro M. Mori H. Sumida M. Matsubara F. Taniani K. Kadono-Okuda K. Kato Y. Mori H. Biochim. Biophys. Acta. 1995; 1260: 245-268Crossref PubMed Scopus (103) Google Scholar, 29Gao Z. Garbers D.L. J. Biol. Chem. 1998; 273: 3415-3421Abstract Full Text Full Text PDF PubMed Scopus (91) Google Scholar, 30Cohen-Salmon M. El-Amraqui A. Leibovici M. Petit C. Proc. Natl. Acad. Sci. U. S. A. 1997; 94: 14450-14455Crossref PubMed Scopus (115) Google Scholar, 31Legan P.K. Rau A. Keen J.F. Richardson G.P. J. Biol. Chem. 1997; 272: 8791-8801Abstract Full Text Full Text PDF PubMed Scopus (200) Google Scholar). Many secreted mucins contain three NH2-terminal D-domains, designated D1, D2, and D3, and some a fourth domain, D4, at the COOH terminus (Fig. 1). A partial D-domain, D′, is between D2 and D3 in all secreted mucins and VWF. Each domain, which contains up to 30 ½Cys, shows significant sequence identity with the other D-domains, especially the half-cystines. Comparisons of the sequences of the D-domains and other ½Cys-rich domains are given as supplemental information (see Supplemental Material). The D1-, D2-, and D3-domains of PSM areN-glycosylated when expressed in COS-7 cells (32Perez-Vilar J. Eckhardt A. DeLuca A.W. Hill R.L. J. Biol. Chem. 1998; 273: 14442-14449Abstract Full Text Full Text PDF PubMed Scopus (59) Google Scholar), but this is not known for other mucins. In PSM and VWF all of the ½Cys in the D1-, D2-, D3-, and CK-domains are thought to form disulfide bonds, some of which are intrachain bonds whereas others are interchain bonds that are involved in assembly of PSM and VWF into multimers (see below). A 240–325-residue domain with 29–33 ½Cys is at the COOH terminus of many mucins (4Gum J.R. Hicks J.W. Toribara N.W. Siddiki B. Kim Y.S. J. Biol. Chem. 1994; 269: 2440-2446Abstract Full Text PDF PubMed Google Scholar, 5Li D. Gallup M. Fan N. Szymkowski D.E. Basbaum C.B. J. Biol. Chem. 1998; 273: 6812-6820Abstract Full Text Full Text PDF PubMed Scopus (156) Google Scholar, 6Desseyn J.L. Aubert J.P. van Seuningen I. Porchet N. Laine A. J. Biol. Chem. 1997; 272: 16873-16883Abstract Full Text Full Text PDF PubMed Scopus (93) Google Scholar, 12Eckhardt A.E. Timpte C.S. DeLuca A.W. Hill R.L. J. Biol. Chem. 1997; 272: 33204-33210Abstract Full Text Full Text PDF PubMed Scopus (48) Google Scholar, 18Joba W. Hoffmann W. J. Biol. Chem. 1997; 272: 1805-1810Abstract Full Text Full Text PDF PubMed Scopus (21) Google Scholar, 33Jiang W. Woitach J.T. Keil R.L. Bhavanandan V.P. Biochem. J. 1998; 331: 193-199Crossref PubMed Scopus (16) Google Scholar, 34Verma M. Davidson E.A. Proc. Natl. Acad. Sci. U. S. A. 1993; 90: 7144-7148Crossref PubMed Scopus (27) Google Scholar) (see Supplemental Material) (Fig. 1). These mucin domains have significant sequence identity with one another and with those at the carboxyl terminus of other proteins (27Sadler J.E. Annu. Rev. Biochem. 1998; 67: 395-424Crossref PubMed Scopus (1109) Google Scholar, 28Kotani E. Yamakawa M. Iwamoto S. Tashiro M. Mori H. Sumida M. Matsubara F. Taniani K. Kadono-Okuda K. Kato Y. Mori H. Biochim. Biophys. Acta. 1995; 1260: 245-268Crossref PubMed Scopus (103) Google Scholar, 30Cohen-Salmon M. El-Amraqui A. Leibovici M. Petit C. Proc. Natl. Acad. Sci. U. S. A. 1997; 94: 14450-14455Crossref PubMed Scopus (115) Google Scholar). Like the D-domains, they are predicted to have globular structures with α-helices and pleated sheets and few or no free thiols (35Perez-Vilar J. Hill R.L. J. Biol. Chem. 1998; 273: 6982-6988Abstract Full Text Full Text PDF PubMed Scopus Google Scholar). They contain oligosaccharides at one or more J. Eckhardt A.E. Hill R.L. J. Biol. Chem. 1996; Full Text Full Text PDF PubMed Scopus Google Scholar). The first residues in this domain have sequence identity with the of but the residues from the COOH terminus have sequence identities with the at the COOH terminus of VWF (27Sadler J.E. Annu. Rev. Biochem. 1998; 67: 395-424Crossref PubMed Scopus (1109) Google Scholar). The CK-domains are to the of proteins that factor and Annu. Rev. Biophys. 1995; PubMed Google Scholar). The CK-domains of VWF and mucins show significant sequence identity to a that in form to in a characterized by and T. A. P. B. C. M. J. Genet. 1993; PubMed Scopus Google Scholar). The the ½Cys that form interchain disulfide bonds between the polypeptide chains of VWF and PSM and other mucins (27Sadler J.E. Annu. Rev. Biochem. 1998; 67: 395-424Crossref PubMed Scopus (1109) Google Scholar, J. Hill R.L. J. Biol. Chem. 1998; 273: 6982-6988Abstract Full Text Full Text PDF PubMed Scopus Google Scholar, J. Eckhardt A.E. Hill R.L. J. Biol. Chem. 1996; Full Text Full Text PDF PubMed Scopus Google Scholar, G. J.F. J. Biochem. 1998; 263: Scopus Google Scholar) (see below). A with sequence identity to those in VWF (27Sadler J.E. Annu. Rev. Biochem. 1998; 67: 395-424Crossref PubMed Scopus (1109) Google Scholar) is found in several mucins (4Gum J.R. Hicks J.W. Toribara N.W. Siddiki B. Kim Y.S. J. Biol. Chem. 1994; 269: 2440-2446Abstract Full Text PDF PubMed Google Scholar, 5Li D. Gallup M. Fan N. Szymkowski D.E. Basbaum C.B. J. Biol. Chem. 1998; 273: 6812-6820Abstract Full Text Full Text PDF PubMed Scopus (156) Google Scholar, 6Desseyn J.L. Aubert J.P. van Seuningen I. Porchet N. Laine A. J. Biol. Chem. 1997; 272: 16873-16883Abstract Full Text Full Text PDF PubMed Scopus (93) Google Scholar, H. Dohrman A.F. Gallup M. Tsuda T. Kai H. Gum J.R. Kim Y.S. Basbaum C.B. J. Biol. Chem. 1994; 269: 17833-17840Abstract Full Text PDF PubMed Google Scholar) (Fig. 1). domains other than the and CK-domains are in a few mucins (Fig. and those in the factor family F. W. J. Biol. Chem. 1992; 267: Full Text PDF PubMed Google Scholar, Annu. Rev. Physiol. 1996; PubMed Scopus Google Scholar) (see Supplemental Material) are in some frog mucins. domains are found in other mucins (8Gum J.R. Ho J.J.L. Pratt W.S. Hicks J.W. Hill A.S. Vinall L.E. Roberton A.M. Swallow D.M. Kim Y.S. J. Biol. Chem. 1992; 267: 26678-26686Abstract Full Text PDF Google Scholar, A.S. M. C.A. Ho S.B. Biochem. J. 1998; 330: PubMed Scopus (48) Google I. G. J. Biochim. Biophys. Acta. 1997; PubMed Scopus Google Scholar). is well known that the molecular of many mucins in the of R.L. A.M. J. N. J. Biol. Chem. Full Text PDF PubMed Google Scholar), that interchain disulfide bonds mucins in a on the of mucins in tissue J. Rev. Biochem. Mol. Biol. 1992; PubMed Scopus Google Scholar) and cells in G. J. Biochem. J. 1994; PubMed Scopus Google Scholar, M. I. A.P. C. Biochem. J. 1996; PubMed Scopus Google Scholar, J. Glycobiology. 1998; PubMed Scopus Google Scholar, N. J. Biol. Chem. 1998; 273: Full Text Full Text PDF PubMed Scopus Google Scholar, N. Biochem. J. 1998; PubMed Scopus Google Scholar) have the of disulfide bonds in the assembly of mucins into The that mucins disulfide-rich domains structurally similar to those in VWF and the that VWF multimers through its disulfide-rich domains (27Sadler J.E. Annu. Rev. Biochem. 1998; 67: 395-424Crossref PubMed Scopus (1109) Google Scholar) a of these domains in mucin the large of mucin polypeptides and their high carbohydrate content of the of for the molecular of mucin it has been to into by of mucin domains in cells by of the recombinant proteins by and and This has been for in PSM (32Perez-Vilar J. Eckhardt A. DeLuca A.W. Hill R.L. J. Biol. Chem. 1998; 273: 14442-14449Abstract Full Text Full Text PDF PubMed Scopus (59) Google Scholar, J. Hill R.L. J. Biol. Chem. 1998; 273: 6982-6988Abstract Full Text Full Text PDF PubMed Scopus Google Scholar, J. Eckhardt A.E. Hill R.L. J. Biol. Chem. 1996; Full Text Full Text PDF PubMed Scopus Google Scholar, J. Hill R.L. J. Biol. Chem. 1998; 273: Scholar), with the that the assembly of domains the assembly of native Thus, as in PSM is thought to form through its and the form multimers through their NH2-terminal is that all mucins structurally to VWF (Fig. in to rat MUC5AC, and MUC6, form multimers similar to those by polypeptide chains of PSM form through their CK-domains their in the (35Perez-Vilar J. Hill R.L. J. Biol. Chem. 1998; 273: 6982-6988Abstract Full Text Full Text PDF PubMed Scopus Google Scholar, J. Eckhardt A.E. Hill R.L. J. Biol. Chem. 1996; Full Text Full Text PDF PubMed Scopus Google Scholar). show that is and with or is not for or are by (32Perez-Vilar J. Eckhardt A. DeLuca A.W. Hill R.L. J. Biol. Chem. 1998; 273: 14442-14449Abstract Full Text Full Text PDF PubMed Scopus (59) Google Scholar, J. Hill R.L. J. Biol. Chem. 1998; 273: 6982-6988Abstract Full Text Full Text PDF PubMed Scopus Google Scholar, J. Eckhardt A.E. Hill R.L. J. Biol. Chem. 1996; Full Text Full Text PDF PubMed Scopus Google Scholar). unglycosylated are secreted and/or into the (32Perez-Vilar J. Eckhardt A. DeLuca A.W. Hill R.L. J. Biol. Chem. 1998; 273: 14442-14449Abstract Full Text Full Text PDF PubMed Scopus (59) Google Scholar). The that a that the has no on and that are oligosaccharides that is to the to the on the of PSM, rat Muc2 also to form through its disulfide-rich domain, which the G. J.F. J. Biochem. 1998; 263: Scopus Google Scholar) (see Supplemental Material). by other types of mucins has not been by of the mucins secreted by cells in M. I. A.P. C. Biochem. J. 1996; PubMed Scopus Google Scholar, J. Glycobiology. 1998; PubMed Scopus Google Scholar, N. J. Biol. Chem. 1998; 273: Full Text Full Text PDF PubMed Scopus Google Scholar, N. Biochem. J. 1998; PubMed Scopus Google Scholar), MUC5AC, and MUC5B and MUC6, appear to form their in the In to PSM, is to be for of rat and The interchain disulfide bonds in PSM have been by (35Perez-Vilar J. Hill R.L. J. Biol. Chem. 1998; 273: 6982-6988Abstract Full Text Full Text PDF PubMed Scopus Google Scholar). Of the ½Cys in the of is on is by of ½Cys at residues and and are in the sequence which is in all mucins and other proteins containing the (see Supplemental Material) and is also for interchain disulfide in VWF (27Sadler J.E. Annu. Rev. Biochem. 1998; 67: 395-424Crossref PubMed Scopus (1109) Google Scholar) and J. Hill R.L. J. Biol. Chem. 1997; 272: Full Text Full Text PDF PubMed Scopus Google Scholar). in PSM in the sequence is also for but the proteins at this are secreted (35Perez-Vilar J. Hill R.L. J. Biol. Chem. 1998; 273: 6982-6988Abstract Full Text Full Text PDF PubMed Scopus Google Scholar), that this sequence may be in of the in the This sequence is also in all mucins, and (Fig. (see Supplemental which to its in the structure of the The oligosaccharides into mucins and as by on MUC2 N. J. Biol. Chem. 1998; 273: Full Text Full Text PDF PubMed Scopus Google Scholar) and MUC5AC N. Biochem. J. 1998; PubMed Scopus Google Scholar) and of PSM J. Y. Eckhardt A.E. Hill R.L. Proc. Natl. Acad. Sci. U. S. A. 1994; PubMed Scopus (115) Google Scholar, M. Eckhardt A.E. J. Hill R.L. J. Biol. Chem. 1988; 263: Full Text PDF PubMed Google Scholar). of PSM when the the the that the and the mucin have been by electron microscopy in the in mucous cells of submaxillary J. Y. Eckhardt A.E. Hill R.L. Proc. Natl. Acad. Sci. U. S. A. 1994; PubMed Scopus (115) Google Scholar, M. Eckhardt A.E. J. Hill R.L. J. Biol. Chem. 1988; 263: Full Text PDF PubMed Google Scholar). Moreover, unglycosylated mucin are in the of the M. Eckhardt A.E. J. Hill R.L. J. Biol. Chem. 1988; 263: Full Text PDF PubMed Google Scholar). cells secreted mucins, cells J. J. Cell Biol. PubMed Scopus Google Scholar), also appear to in although in cell is found to in the G. J. Biochem. J. 1994; PubMed Scopus Google Scholar, J. J. A. J. Biol. Chem. 1991; Full Text PDF PubMed Google Scholar). The of the of oligosaccharides in secreted mucins in the and the for and of the oligosaccharides are P. G. Rev. Biochem. Mol. Biol. 1998; PubMed Scopus Google Scholar). in COS-7 cells of the three D-domains of PSM has shown that these domains in of interchain disulfide bonds between to high molecular multimers of mucin (32Perez-Vilar J. Eckhardt A. DeLuca A.W. Hill R.L. J. Biol. Chem. 1998; 273: 14442-14449Abstract Full Text Full Text PDF PubMed Scopus (59) Google Scholar). from in several which the that multimers form in the that the of as and also but not (32Perez-Vilar J. Eckhardt A. DeLuca A.W. Hill R.L. J. Biol. Chem. 1998; 273: 14442-14449Abstract Full Text Full Text PDF PubMed Scopus (59) Google Scholar). a of the that the at a also These suggest that the interchain disulfide bonds that to multimers are at a in the through ½Cys residues in the The molecular weights of the multimers cannot be by they are large they not the with a of were when the three D-domains were expressed (32Perez-Vilar J. Eckhardt A. DeLuca A.W. Hill R.L. J. Biol. Chem. 1998; 273: 14442-14449Abstract Full Text Full Text PDF PubMed Scopus (59) Google Scholar), that a in the of is of Such multimers are structures as in PSM containing no glycosylated domains is secreted from COS-7 cells as and multimers and that VWF not all are to multimers (32Perez-Vilar J. Eckhardt A. DeLuca A.W. Hill R.L. J. Biol. Chem. 1998; 273: 14442-14449Abstract Full Text Full Text PDF PubMed Scopus (59) Google Scholar). in the of PSM has been found by to be a for one of the interchain disulfide bonds in mucin multimers J. Hill R.L. J. Biol. Chem. 1998; 273: Scholar). is in the sequence which is highly in secreted mucins (Fig. (see Supplemental and the ½Cys in VWF is for its (27Sadler J.E. Annu. Rev. Biochem. 1998; 67: 395-424Crossref PubMed Scopus (1109) Google Scholar). In which is to form interchain disulfide bonds in VWF (27Sadler J.E. Annu. Rev. Biochem. 1998; 67: 395-424Crossref PubMed Scopus (1109) Google Scholar), not form bonds in PSM J. Hill R.L. J. Biol. Chem. 1998; 273: Scholar). The other in the D-domains that form interchain disulfide bonds are not VWF multimers are from which is by a at in the sequence in the (27Sadler J.E. Annu. Rev. Biochem. 1998; 67: 395-424Crossref PubMed Scopus (1109) Google Scholar). The contains the and and is for although is may not be for mucin the of mucins not contain the sequence for of The that the D-domains of PSM are not when expressed in COS-7 or cells (32Perez-Vilar J. Eckhardt A. DeLuca A.W. Hill R.L. J. Biol. Chem. 1998; 273: 14442-14449Abstract Full Text Full Text PDF PubMed Scopus (59) Google Scholar) is with the lack of the in the of PSM (see Supplemental Material). some of mucins is as by that in the of MUC2 A. J.R. G. S. Swallow D.M. I. J. Biol. Chem. 1999; Full Text Full Text PDF PubMed Scopus Google Scholar), MUC5B C. J.R. I. Biochem. J. 1998; PubMed Scopus Google Scholar), and rat Muc2 J.F. J. 1996; PubMed Scopus Google Scholar), although a for in mucin assembly is Moreover, of the mucins not The that PSM contains multimers that be on electron microscopy of mucins. is that mucins form J. Rev. Biochem. Mol. Biol. 1992; PubMed Scopus Google Scholar), which has been by electron microscopy (24Eckhardt A.E. Timpte C.S. Abernethy J.L. Toumadje A. Johnson Jr., W.C. Hill R.L. J. Biol. Chem. 1987; 262: 11339-11344Abstract Full Text PDF PubMed Google Scholar). Moreover, VWF through its and the form multimers through their (27Sadler J.E. Annu. Rev. Biochem. 1998; 67: 395-424Crossref PubMed Scopus (1109) Google Scholar). is that some mucins with disulfide-rich domains not form multimers in the for mucins are highly to A.E. Timpte C.S. Abernethy J.L. Y. Hill R.L. J. Biol. Chem. 1991; Full Text PDF PubMed Google Scholar, J.F. Biochem. J. 1998; 331: PubMed Scopus (21) Google Scholar), and electron be on well characterized Of is a structures for MUC5B in of M. Richardson T. Biochem. J. 1999; 338: PubMed Scopus (91) Google Scholar). Nevertheless, of mucin assembly are by on cells MUC2 but molecular are N. J. Biol. Chem. 1998; 273: Full Text Full Text PDF PubMed Scopus Google Scholar). the are assembled in the by a These are by chemical bonds of N. J. Biol. Chem. 1998; 273: Full Text Full Text PDF PubMed Scopus Google Scholar). is not known the are although MUC2 in the is thought to be part of glycoprotein A. J.R. G. S. Swallow D.M. I. J. Biol. Chem. 1999; Full Text Full Text PDF PubMed Scopus Google Scholar). suggest that MUC2 is assembled into large M. I. A.P. C. Biochem. J. 1996; PubMed Scopus Google N. A. C. Glycobiology. 1999; PubMed Scopus Google Scholar). MUC5AC J.R. I. Biochem. J. 1996; PubMed Scopus Google Scholar) and MUC5B C. J.R. I. Biochem. J. 1998; PubMed Scopus Google Scholar) are large mucins by disulfide described the assembly of PSM and VWF (27Sadler J.E. Annu. Rev. Biochem. 1998; 67: 395-424Crossref PubMed Scopus (1109) Google Scholar) in the and in the The molecular that this are not but the NH2-terminal D-domains and the in the and D3-domains to J. Hill R.L. J. Biol. Chem. 1998; 273: Scholar). the and the and the or the of PSM expressed mucin in the of that the three domains be to at the of the and the of the two ½Cys by in the in the of multimers in the of J. Hill R.L. J. Biol. Chem. 1998; 273: Scholar). Thus, the in the of mucin in the of the and of the two ½Cys by in the in the the of of multimers J. Hill R.L. J. Biol. Chem. 1998; 273: Scholar). This suggests that at in the the in the of VWF also the in the (27Sadler J.E. Annu. Rev. Biochem. 1998; 67: 395-424Crossref PubMed Scopus (1109) Google Scholar), although a for the in the has not been VWF has another in the that is also for its assembly (27Sadler J.E. Annu. Rev. Biochem. 1998; 67: 395-424Crossref PubMed Scopus (1109) Google Scholar). among the mucins structurally to MUC5AC and MUC5B have in their (see Supplemental Material). The of the but of the that similar are in the of proteins involved in of disulfide bonds as disulfide (Fig. the these have a in of disulfide bonds in mucins. has been in of the structure and assembly of mucins, but much for the of the mucin family be and their structures and of assembly into multimers The of to form the many disulfide bonds in the globular domains be and the of in of these domains also be determined. The molecular for the of mucins be These of will be to into the of mucins. with