bluuebunny/crossref_metadata_2025 huggingface dataset:
Cleavage of four avian sarcoma virus polyproteins with virion protease p15 removes gag sequences and yields large fragments that function as tyrosine phosphoacceptors in vitro.
Authors: Ghysdael J , Neil J C , Vogt P K
Published: May 2006

The transformation-specific polyproteins of avian sarcoma viruses PRCII, PRCII-p, Fujinami sarcoma virus (FSV), and Esh sarcoma virus (ESV) consist of two domains, one derived from a partial viral gag gene and the other representing an apparently cell-derived insert in the defective viral genome. These gag-linked proteins were cleaved with retrovirion protease p15. Cleavage of PRCII-p polyprotein P170, P105 of PRCII, and P140 of FSV occurred within the gag domain and generated fragments of Mr 130,000, 70,000, and 115,000, respectively, containing all of the transformation-specific sequences linked to a remnant of the original gag sequences. ESV P80 was cleaved inside the transformation-specific domain, yielding a Mr 35,000--38,000 fragment from the NH2-terminal half of the molecule consisting of the entire gag portion and some no-gag sequences and a Mr 48,000 fragment containing most of the transformation-specific sequences. The tyrosine phosphorylation sites of the polyproteins were found in every case in the transformation-specific fragments. The major serine phosphorylation site of ESV P80 was found to reside in the Mr 35,000--38,000 gag-containing fragment, probably within the transformation-specific sequences of that cleavage product. Removal of all of the gag domain of ESV P80 or most of the gag domain in PRCII-p P170, PRCII P105, and FSV P140 does not affect their ability to be phosphorylated by the polyprotein-associated tyrosine-specific protein kinase activities. This observation suggests that the gag sequences of the polyproteins of classes II (PRCII-p, PRCII, and FSV) and III (ESV) avian sarcoma viruses may not be required for this enzymatic function, which appears to be of importance in transformation.
Population structure of multilocus associations
Authors: Brown A. H. D. , Feldman M. W.
Published: May 2006

A method of analysis is presented whereby the structure of multilocus associations among and within several populations can be partitioned into its components. The components are measured by their contributions to the variance in the number of heterozygous loci in two randomly chosen gametes. The singlelocus components are the average and the variation among populations in gene diversity and the variance among populations in allele frequency. The two-locus components include the mean and variance of disequilibria, the covariance of allele frequencies over populations, and various interactions. When applied to allozyme data from populations of wild (
Hordeum spontaneum
) and cultivated barley (
H. vulgare
), the analysis highlighted the repetitive pattern of the multilocus associations in the composite crosses whereas it emphasized the regionally localized and geographically variable pattern present in the natural populations of the wild species. The analysis is flexible and applicable to multilocus gametic data from any set of populations, without regard to the number of alleles per locus or the reproductive method of the organism.
Energy-filtered transmission electron microscopy of ferritin.
Authors: Shuman H , Somlyo A P
Published: May 2006

The focusing properties of a magnetic-sector spectrometer are shown to be suitable for forming high-spatial-resolution, energy-filtered transmission electron microscope images. Filtered images of ferritin molecules by using electrons scattered from the characteristic iron M2,3 and carbon K absorption edges clearly distinguish the 75-A iron core and 120-A protein shell. The minimum detectable mass is estimated to be 0.84 X 10(-20) g for Fe for an electron dose of 18 C/cm2 and 99% confidence.
Identification and characterization of the ecdysterone receptor in
Drosophila melanogaster
by photoaffinity labeling
Authors: Schaltmann Karin , Pongs Olaf
Published: May 2006

Salivary glands of third-instar larvae of
Drosophila melanogaster
as well as
Drosophila
K
c
tissue culture cells have been irradiated in the presence of ecdysterone. Irradiation covalently links ecdysterone to a single cellular protein, which is similar, if not identical, in salivary glands and in K
c
cells. This protein has a molecular weight of 130,000 and it has the characteristics of a typical hormone-receptor molecule in terms of hormone-binding properties, translocation into the nucleus, and sedimentation characteristics. The yield of the photoinduced bonding of ecdysterone to receptor protein is around 15%. Ponasterone A competed with ecdysterone for the bonding. Also, ponasterone A itself reacted upon photoactivation with the β-ecdysterone receptor protein in
Drosophila
tissue culture cells. We have previously shown that ecdysterone can be bonded upon irradiation to specific hormone-controlled puffs of polytene chromosomes of
D. melanogaster
third-instar larvae [Gronemeyer, H. & Pongs, O. (1980)
Proc. Natl. Acad. Sci. USA
77, 2108-2112]. Because we have now identified the molecular target of the ecdysterone photoreaction, these data show that a hormone-receptor complex translocates to the nucleus and directly binds to the genes, which are under hormonal control. A quantitative assay of hormone-receptor complex in K
c
cells before and after hormone stimulation showed that ecdysterone does not regulate the synthesis and the available amount of its receptor. It was also observed that the translocated hormone-receptor complex resides in the nucleus as long as the hormone is present in the tissue culture medium.
Assembly of new histones into nucleosomes and their distribution in replicating chromatin.
Authors: Russev G , Hancock R
Published: May 2006

We studied the assembly of new histones into nucleosomes and their distribution in replicating chromatin in growing P815 mouse cells. New histones and new DNA were density-labeled with 13C, 15N, 2H-substituted amino acids together with [3H]arginine or with 5-iododeoxyuridine and [3H]thymidine, respectively, for 1 hr (approximately 20% of S phase). Mono- di-, tri-, tetra- and larger oligonucleosomes were isolated by sucrose gradient centrifugation of micrococcal nuclease-digested chromatin, and their density distribution was analyzed, without fixation, in metrizamide/triethanolamine density gradients [Russev, G. and Tsanev, R. (1976) Nucleic Acids Res. 3, 697-707] in which mono- and oligonucleosomes containing dense amino acids or 5-iododeoxyuridine separate from the corresponding normal nucleosomes. Under these conditions, approximately 74% of the new histones are found in nucleosomes on newly replicated DNA, and the remainder are on unreplicated DNA. The majority of new histones form entirely new nucleosomes; a minor fraction may form hybrid nucleosomes that also contain preexisting histones. New nucleosomes are distributed to both new daughter DNA molecules with approximately equal probability, and our evidence suggests, but does not prove, that they are distributed in a random manner along new DNA.
Recombinational bypass of pyrimidine dimers promoted by the recA protein of Escherichia coli.
Authors: Livneh Z , Lehman I R
Published: May 2006

recA protein, in the presence of single-stranded DNA binding protein and ATP, promotes the complete exchange of strands between circular single-stranded DNA containing pyrimidine dimers and a homologous linear duplex, converting the pyrimidine dimer-containing single-stranded DNA to a circular duplex. Bypass of a pyrimidine dimer during the branch-migration phase of the reaction requires approximately 20 seconds, a rate 1/50th of that in the absence of the dimer. The circular duplex product is specifically incised by the pyrimidine dimer-specific T4 endonuclease V, and the resulting 3' hydroxyl termini can serve as primers for deoxynucleotide polymerization by DNA polymerase I. These findings indicate that recA protein serves a direct role in recombinational repair and demonstrate that the pyrimidine dimers that have been bypassed can be processed by enzymes of the excision-repair pathway.
P1 site-specific recombination: nucleotide sequence of the recombining sites.
Authors: Hoess R H , Ziese M , Sternberg N
Published: May 2006

Site-specific recombination between molecules of bacteriophage P1 DNA occurs at sites called loxP and requires the action of a protein that is the product of the P1 cre gene. Although recombination between two loxP sites is very efficient, recombination between loxP and a unique site in the bacterial chromosome (loxB) is inefficient and generates two hybrid lox sites called loxR and loxL. We present here the nucleotide sequences of all four lox sites. Analysis of these sequences indicates that (i) a region of extensive homology is not present at the loxP X loxB crossover point, in contrast to the 15-base pair common-core sequence in the bacteriophage lambda att sites, and (ii) the sites contain a region of dyad symmetry with 8- to 13-base pair inverted repeats separated by an 8- to 9-base pair sequence. The loxP X loxB crossover point falls in the sequence that separates the inverted repeats, and deletions that remove either the left or the right inverted repeat of loxP inactivate the site. These two observations are consistent with the conclusion that the region of dyad symmetry is important in los recombination. We have shown further that the loxP X loxP crossover point occurs in a 63-base pair sequence containing the loxP X loxB crossover point, suggesting that, despite the great difference in efficiencies of the two reactions, the crossover points may occur at the same place in both. Explanations for the different recombination properties of the various lox sites are discussed.
Protein complexes from active replicative fractions associate in vitro with the replication origins of yeast 2-micrometers DNA plasmid.
Authors: Jazwinski S M , Edelman G M
Published: May 2006

In a search for a replication complex, the activity that replicates the 2-micrometers yeast DNA plasmid in vitro was isolated in a high molecular weight form (Mr approximately 2 X 10(6) by gel filtration and rate-zonal sedimentation from extracts prepared from cells of the budding yeast Saccharomyces. When obtained from cells in late logarithmic cultures this material or "complex" was labile compared to that from early logarithmic cultures, and it did not survive as a complex after ammonium sulfate precipitation. This suggests that, as cultures approach stationary phase and cells cease growth, the association of its protein constituents may be altered. A chimera of 2-micrometers DNA inserted into the plasmid pBR322 was used to test for binding of components of the complex. After a brief incubation of the chimera in vitro with the high molecular weight material containing replicating activity, a protein "knob" was found associated with the 2-micrometers DNA as shown by electron microscopy. This association was not random but was limited to two positions on the plasmid. In the same series of experiments, the in vitro origins of 2-micrometers plasmid replication were also mapped. Two origins were found, consistent in position with those that have been identified in vivo. Molecules utilizing both origins simultaneously in vitro were not observed, and replication in vitro was bidirectional. The location of the origins corresponded to the positions at which the protein knobs associated with 2-micrometers DNA. This and the fact that no replicative intermediates with associated complexes were detected raises the possibility that a specific protein complex may be involved in initiation of DNA replication.
Evidence for a role of calmodulin in serum stimulation of Na+ influx in human fibroblasts.
Authors: Owen N E , Villereal M L
Published: May 2006

Sodium influx in serum-deprived human diploid fibroblasts can be stimulated by addition of serum (5-fold) or the divalent cation ionophore A23187 (3-fold). The possible involvement of calmodulin in serum or A23187 stimulation of Na+ influx has been investigated by using six psychoactive agents that are known to bind calmodulin and inhibit calmodulin-sensitive enzymes. Each agent inhibited serum- and A23187-stimulated Na+ influx in a dose-dependent manner. Furthermore, the K1 for inhibition of serum-stimulated Na+ influx correlates directly with the Ca2+-specific calmodulin binding previously determined in a cell-free system [Levin, R. M. & Weiss, B. (1979) J. Pharmacol. Exp, Ther. 208, 454--459]. None of the agents tested had any effect on the serum-insensitive component of net Na+ influx in these cells. These data support the concept that serum and A23187 stimulate Na+ influx in human diploid fibroblasts via an increase of intracellular Ca2+ and a subsequent calmodulin-mediated activation of the amiloride-sensitive transport pathway.
Monoclonal antibody specific for myelin glycoprotein P0: derivation and characterization.
Authors: Franko M C , Koski C L , Gibbs C J , McFarlin D E , Gajdusek D C
Published: May 2006

A monoclonal antibody was produced against the major structural glycoprotein (P0) of human peripheral nervous system myelin. The hybridomas were generated by fusion of mouse myeloma line NS-1 with spleen cells of C3H mice immunized with purified human peripheral nervous system myelin. Hybridomas were screened by a two-step solid-phase radioimmunoassay, with P0 adsorbed on microtiter plates and with addition of 125I-labeled rabbit anti-mouse IgG as the second step. One derived clone, designated 41G10, bound P0 in the radioimmunoassay 4-fold over the background value obtained by using bovine serum albumin as the negative control antigen. Clone 41G10 was shown by immunofluorescence to bind to frozen sections of human intercostal nerve. Diffuse fluorescent staining occurred uniformly over the entire myelin sheath. The cylindrical axons were unstained. The same pattern of immunofluorescence was noted on rat, hamster, mouse, and rabbit sciatic nerve. Immunofluorescence was abolished when 41G10 was absorbed by P0. The monoclonal antibody 41G10 was absorbed by P0. The monoclonal antibody 41G10 is of the IgM class and activated complement in the presence of myelin vesicles or P0 liposomes but not in their absence.
Replication and expression of a bacterial--mitochondrial hybrid plasmid in the fungus Podospora anserina.
Authors: Stahl U , Tudzynski P , Kück U , Esser K
Published: May 2006

Hybrid plasmids consisting of the bacterial plasmid pBR322 and plasmid-like DNA (pl DNA) sequences from the fungus Podospora anserina are not only able to replicate in Escherichia coli but also in the fungus. This was proved by both biophysical and biological evidence involving buoyant density profiles, DNA.DNA hybridization, and restriction analysis--all confirming that pl DNA behaves as a true plasmid. During its amplification in P. anserina, the hybrid plasmid does not lose its prokaryotic coding capacity as shown after retransfer and subsequent cloning in E. coli. P. anserina is able to express both the eukaryotic and the prokaryotic genetic information of the hybrid plasmid because the occurrence of senescence and the production of beta-lactamase could be shown in experiments involving specific hybrid plasmids. In the same systems, it was possible to demonstrate that a hybrid plasmid containing, instead of pl DNA, a pl DNA homologous region of native mtDNA also could function as a true plasmid. This hybrid plasmid contained about 25% of the genetic information pl DNA, which corresponds to about 6% of the genetic information of mtDNA. Thus, the data show that hybrid plasmids may be used to shuttle genetic information between P. anserina and E. coli. hence, through the use of a mtDNA replicon, as evidenced by the pl DNA of P. anserina, another pathway in genetic engineering is established.
In vitro replication of recombinant plasmids carrying chromosomal segments of Xenopus laevis.
Authors: Hiraga S , Sudo T , Yoshida M , Kubota H , Ueyama H
Published: May 2006

Recombinant plasmids carrying a segment of Xenopus laevis chromosomal DNA were constructed with plasmid pBR322 as the vector. A recombinant plasmid pXY65 carrying a 3.2-kilobase BamHI segment of the chromosome of X. laevis has been found to contain a repetitive sequence dispersed throughout the X. laevis chromosomes. This plasmid initiated replication in vitro when the supercoiled circular molecules were incubated in a replication system. The other recombinant plasmids tested and the pBR322 vector were not replicated. Electron microscopic analysis of the replicative intermediates showed that the replication was initiated at a specific site in the 3.2-kilobase BamHI segment of pXY65 and that the replication usually proceeded bidirectionally. Analysis of the reaction products by centrifugation in alkaline sucrose gradients indicated that short pieces were synthesized in the in vitro replication system. DNA synthesis was inhibited in vitro by the addition of aphidicolin and by omission of dNTPs. These results indicate that the X. laevis segment cloned in pXY65 contains a site capable of initiating replication in vitro.
Catalytic mechanism of glycogen phosphorylase: pyridoxal(5')diphospho(1)-alpha-D-glucose as a transition-state analogue.
Authors: Takagi M , Fukui T , Shimomura S
Published: May 2006

Pyridoxal(5')diphospho(1)-alpha-D-glucose was used to reconstitute glycogen phosphorylase beta (1,4-alpha-D-glucan:orthophosphate alpha-D-glucosyltransferase, EC 2.4.1.1) from rabbit muscle, replacing the natural pyridoxal 5'-phosphate coenzyme. Incubation of the reconstituted enzyme alone resulted in the gradual cleavage of the synthetic cofactor to pyridoxal 5'-phosphate, which caused slow reactivation of the enzyme. The addition of maltopentaose or glycogen altered the mode of cleavage; the cofactor was rapidly decomposed to pyridoxal 5'-diphosphate. The radioactive glucose moiety released from pyridoxal(5')diphospho(1)-alpha-D-[14C]glucose was incorporated into the outer chain of glycogen, forming an alpha-1,4-glucosidic linkage. These results show that the glucosyl transfer reaction discovered mimics the normal catalysis of this enzyme, and they strongly support the catalytic mechanism in which the coenzyme phosphate acts as a catalyst by direct interaction with the phosphate of the substrate, forming the pyrophosphate-like transition intermediate.
Inhibition of glucose transport in human erythrocytes by cytochalasins: A model based on diffraction studies.
Authors: Griffin J F , Rampal A L , Jung C Y
Published: May 2006

On the basis of details of the three-dimensional structures of beta-D-glucose and of cytochalasins, either previously published or reported here (cytochalasin A), we propose a model to explain the observed difference in activity of cytochalasins in the inhibition of glucose transport. In our model cytochalasin B binds to the glucose carrier through hydrogen bonds at N2 (donates), O7 (accepts), and O23 (accepts) analogous to O6, O3, and O1, respectively, on beta-D-glucose. The hydrophobic region from C13 to C19 is also essential in binding and appears to act as an anchor in a hydrophobic domain of the glucose carrier. The presence of hydrophilic groups in this essential hydrophobic region accounts, at least in part, for the inactivity of the other cytochalasins in the series.
Down-regulation of the hexose transport system: metabolic basis studied with a fibroblast mutant lacking phosphoglucose isomerase.
Authors: Ullrey D B , Franchi A , Pouyssegur J , Kalckar H M
Published: May 2006

Down-regulation ("curb") of hexose transport in Chinese hamster lung fibroblasts has been studied in a metabolic mutant highly defective in phosphoglucose isomerase (PGI; glucosephosphate isomerase; D-glucose-6-phosphate ketol-isomerase, EC 5.3.1.9). In the parental strain (PGI+) glucose as well as glucosamine and mannose were able to elicit a curb of the hexose transport system. In the PGI mutant, only glucose was able to mediate a transport curb. The inability of glucosamine and mannose to promote a transport curb in the PGI strain must be ascribed to the fact that the 6-esters of these aldohexoses are converted by their own specific deaminase and isomerase to fructose 6-phosphate, which initiates the pyruvate-tricarboxylate energy-yielding pathway but cannot be converted to glucose 6-phosphate in the mutant. The latter ester can be metabolized, but its metabolism in the mutant is confined to the pentose shunt. It is shown that inhibitors such as 2,4-dinitrophenol and malonate exert only slight inhibition of the pentose shunt yet release the glucose-mediated curb elicited by glucose and glucosamine in the parental PGI+ strain and also the glucose transport curb persisting in the PGI mutant.
Herpesvirus-specific RNA and protein in carcinoma of the uterine cervix.
Authors: McDougall J K , Crum C P , Fenoglio C M , Goldstein L C , Galloway D A
Published: May 2006

Cloned probes of herpes simplex virus type 2 DNA were used in cytological hybridization experiments to detect herpesvirus RNA transcripts in the neoplastic cells of tumors of the uterine cervix. Virus-specific RNA was shown to represent transcription of limited regions of the genome, of which one is known to code for a DNA-binding protein that can be found by immunoperoxidase staining in the neoplastic cells of these tumors and has also been detected in cells transformed in vitro by this virus.
Lower bounds for the life-span of solutions of nonlinear wave equations in three dimensions
Authors: John Fritz
Published: May 2006

The paper deals with strict solutions
u
(
x,t
) =
u
(
x
1
,
x
2
,
x
3
,
t
) of an equation [Formula: see text] where
Du
is the set of four first derivatives of
u
. For given initial values
u
(
x
,0) = ε
F
(
x
),

u
t

(
x
,0) = ε
G
(
x
), the life span
T
(ε) is defined as the supremum of all
t
to which the local solution can be extended for all
x
. Blowup in finite time corresponds to
T
(ε) < ∞. Examples show that this can occur for arbitrarily small ε. On the other hand,
T
(ε) must at least be very large for small ε. By assuming that

a
ik
,F,G

[unk]
C
∞
, that

a
ik

(0) = 0, and that
F,G
have compact support, it is shown that [Formula: see text] for every
N
. This result had been established previously only for
N
< 4.
Equilibrium cooperative binding of calcium and protons by sarcoplasmic reticulum ATPase.
Authors: Hill T L , Inesi G
Published: May 2006

The cooperative equilibrium binding of Ca2+ by sarcoplasmic reticulum ATPase, as modulated by pH, is analyzed by statistical mechanical treatment of a theoretical model. The model consists of four equivalent subunits, in the form of a square, with nearest-neighbor interactions. Each subunit has one site for binding of one Ca2+ or one proton, but not both. Binding of either ligand on a subunit induces a conformational change in the subunit that alters its interaction with its two neighbors. The model gives good agreement with experimental binding data. It should prove useful as a starting point in the analysis of steady-state ATPase activity as a function of Ca2+ and H+ concentrations.
Asymmetric replication of duck hepatitis B virus DNA in liver cells: Free minus-strand DNA.
Authors: Mason W S , Aldrich C , Summers J , Taylor J M
Published: May 2006

In order to study the replication of the DNA genome of duck hepatitis B virus, an avian virus related to human hepatitis B virus, we have characterized viral DNAs present in the livers of viremic ducks by agarose gel electrophoresis and the Southern blot procedure. In addition to relaxed circular DNA similar to virion DNA, livers contained a heterogeneous population of rapidly migrating species. The conformation of the rapidly migrating species was markedly sensitive to salt, suggesting that these species were largely single stranded. The largest major rapidly migrating species was shown to have an electrophoretic mobility that was insensitive to preheating of the DNA to 100 degrees C and was similar to that of denatured virus DNA 3 kilobases long, suggesting that this DNA was a single-stranded copy of the entire virus genome. Hybridization with strand-specific probes demonstrated that this 3-kilobase species, as well as more rapidly migrating DNAs, were predominantly minus strands.
Model for the participation of quasi-palindromic DNA sequences in frameshift mutation.
Authors: Ripley L S
Published: May 2006

A model is described for the templated production of frameshift and base-substitution mutations mediated through aberrant DNA structures arising as a consequence of quasi-palindromic DNA sequences. Two general mechanisms are considered. One evokes the formation and processing of imperfect DNA secondary structures (hairpins) for the production of mutations. The other evokes a "strand switch" during DNA synthesis which, in a manner unique to quasi-palindromic sequences, may be resolved to produce frameshift or base-substitution mutations, or both. It is the unique combination of symmetrical and asymmetrical elements of the quasi-palindromic sequence itself that provides the basis for both models. Through the mechanisms described, the symmetrical elements permit unusually paired DNA substrates, and the asymmetrical elements permit templated insertions, deletions, and base substitutions. The model predicts a class of mutations--simultaneously frameshifts and base substitutions--whose sequences can be predicted from a local quasi-palindromic sequence. This prediction appears to be met by a significant fraction (more than 15%) of frameshift mutations in the iso-1-cytochrome c gene of Saccharomyces cerevisiae.
Activation of human erythrocyte Ca2+-dependent Mg2+-activated ATPase by calmodulin and calcium: quantitative analysis.
Authors: Cox J A , Comte M , Stein E A
Published: May 2006

The effect of Ca2+ and calmodulin on (CaM) on the activation of Ca2+-dependent Mg2+-activated ATPase (Ca2+,Mg2+-ATPase; ATP phosphohydrolase, EC 3.6.1.3) has been carried out because of the finding that the CaM dependence of the activation varies with the concentration of free Ca2+, similarly to brain phosphodiesterase and adenylate cyclase. The study was carried out in the absence of chelating agents because they strongly interfere in the enzyme kinetics. Three main conclusions can be drawn (i) CaM-Ca3 and CaM-Ca4 together are the biochemically active species in vitro. (ii) These species bind in a non-cooperative way to the CaM-binding site of the enzyme with a dissociation constant of 6 x 10(-10) M or 1.1 x 10(-8) M, depending on whether Ca2+ saturates the substrate binding site of the enzyme or not. (iii) The binding of CaM-Ca3 to the enzyme lowers the dissociation constant of the enzyme for Ca2+ at the substrate binding site from 51.5 to 2.8 microM. Contrary to general belief, CaM does not induce pronounced positive cooperativity in the binding of Ca2+ to the enzyme. Such a cooperativity is seen only when the enzyme is incompletely saturated with the activator, but it disappears in the presence of saturating concentrations of CaM-Ca3. The rate equation proposed here accurately predicts the extent of enzyme activation over a wide range of Ca2+ and CaM concentration. In healthy erythrocytes the concentrations of Ca2+ and CaM are such that the Ca pump works with a minimal dissipation of energy, but a small increase in the intracellular Ca2+ concentration leads to a strong amplification of the pumping activity.
Immune response to synthetic peptide analogues of hepatitis B surface antigen specific for the a determinant.
Authors: Bhatnagar P K , Papas E , Blum H E , Milich D R , Nitecki D , Karels M J , Vyas G N
Published: May 2006

Recovery from a natural infection with hepatitis B virus or vaccination with purified envelope protein leads to production of antibodies against the hepatitis B surface antigen (HBsAg). Such physiologic response in man is generally directed against the a determinant of HBsAg common to all serotypes of the virus. To define the immunochemical specificity of this determinant, the secondary structure of HBsAg was derived from its sequence of 226 amino acids. Hydrophilic stretches expected to contain the antigenic determinants were located between residues 32 and 76 and between residues 110 and 156. Loss of the antigenic activity after chemical modification of lysine residues of HBsAg indicated their critical importance in antigenicity. Because all lysines are located between residues 121 and 160, we selected this region for localization of HbsAg determinants. Solid-phase synthesis was used to prepare seven peptide analogues of HBsAg (PsAs): 122-137, 128-134, 139-147, 139-158, 140-158, 145-158, and 150-158. For experimental immunization of rabbits the synthetic peptides were coupled to keyhole limpet hemocyanin. We studied the antigenicity of each peptide analogue by serologic neutralization of human antibodies specific for the a determinant of HBsAg. Analogues 139-147, 139-158, and 140-158 showed antigenicity as well as function of anti-HBsAg. The rabbit antibodies were inhibited with each of the three peptide analogues and all serotypes of natural HbSag, having only the a determinant in common. These results indicate that the nonapeptide sequence 139-147 represents the total or an essential part of the a determinant of HBsAg.
The bee's map of the e-vector pattern in the sky
Authors: Rossel Samuel , Wehner Rüdiger
Published: May 2006

It has long been known that bees can use the pattern of polarized light in the sky as a compass cue even if they can see only a small part of the whole pattern. How they solve this problem has remained enigmatic. Here we show that the bees rely on a generalized celestial map that is used invariably throughout the day. We reconstruct this map by analyzing the navigation errors made by bees to which single e-vectors are displayed. In addition, we demonstrate how the bee's celestial map can be derived from the e-vector patterns in the sky.
alpha-Helix dipole model and electrostatic stabilization of 4-alpha-helical proteins.
Authors: Sheridan R P , Levy R M , Salemme F R
Published: May 2006

A simple dipole model is developed for estimation of the electrostatic interaction energy between alpha-helices in proteins. This model is used to estimate the electrostatic stabilization in a recurrent protein tertiary structural motif, an array of four closely packed alpha-helices. It is found that, for the proteins examined (cytochrome c', hemerythrin, myohemerythrin, cytochrome b562, and a T4 phage lysozyme domain), their common antiparallel arrangement of adjacent helices confers a stabilization of 5--7 kcal/mol (1 cal = 4.18 J). In contrast, a similarly packed array of parallel helices is relatively destabilized by 20 kcal/mol. These results show that helix-dipole interactions are important in the stabilization of this structural motif. These effects are discussed both in the context of folding pathways for 4-alpha-helical proteins and the stabilization of the higher aggregates.
Adsorptive endocytosis of fibrin monomer by macrophages: evidence of a receptor for the amino terminus of the fibrin alpha chain.
Authors: Gonda S R , Shainoff J R
Published: May 2006

Human fibrinogen and fibrin monomer were labeled with heme-octapeptide for cytochemical examination of their interaction with rabbit peritoneal macrophages in vitro. Upon short exposure to labeled fibrin monomer in solution with unlabeled fibrinogen, the cells became covered with surface-adsorbed monomer in nonaggregated form and having a characteristic trinodular shape. Within 30 min, the adsorbed monomer became fully internalized by vesicular uptake, with much of it being incorporated into lysosomal bodies. A concomitant loss of adsorptive capacity of the cell surface for uptake of more monomer accompanied the internalization. By contrast, labeled fibrinogen was not adsorbed by the macrophage surface. Some internalization of fibrinogen by passive fluid-phase pinocytosis was evident, but it was not accompanied by loss of absorptive capacity for fibrin monomer. The active uptake of monomer may have depended on binding to the amino terminus that is blocked by fibrinopeptide A in fibrinogen, because addition of synthetic peptide corresponding to the terminal Gly-Pro-Arg segment inhibited both the adsorption and the internalization of monomer.
Cloned endogenous retroviral sequences from human DNA.
Authors: Bonner T I , O'Connell C , Cohen M
Published: May 2006

We have screened a human DNA library using as probe a chimpanzee sequence that contains homology to the polymerase gene of the endogenous baboon virus. One set of overlapping clones spans about 20 kilobases and contains regions of DNA sequence homology to the gag p30, gag p15, and polymerase genes of Moloney murine leukemia virus. Furthermore, the spacings are the same as in Moloney virus between these sequences and a 480-nucleotide region that has the structural characteristics of a 3' copy of the long terminal repeat sequence. Hybridization of the cloned DNA to restriction digests of human DNA indicates that the human genome contains only two copies closely related to the sequence and approximately equal to 10 less closely related copies. This retroviral sequence appears to have been in its present chromosomal location prior to the divergence of man and chimpanzee because the human and chimpanzee clones have 3--4 kilobases of identical 3' flanking sequence.
Protection against a nonimmunogenic mouse leukemia by an immunogenic variant obtained by mutagenesis.
Authors: Van Pel A , Boon T
Published: May 2006

The nonimmunogenic thymic leukemia TH, obtained in mouse strain CBA/Ht, was adapted to culture. By in vitro treatment of a clonal TH cell line with the mutagen N-methyl-N'-nitro-N-nitrosoguanidine, stable variant cell clones (tum-) were obtained that elicited a rejection response in syngeneic mice. Mice that had rejected a tum- variant were partially protected against a challenge with the original tumor. When spleen cells of these animals were restimulated in vitro, cytolytic T cells were obtained that were directed against an antigen present on the original tumor. The existence of these cytolytic effectors was confirmed by clonal analysis of the cytolytic response. No immune protection against TH was observed in mice that had been immunized with irradiated cells of the original TH tumor. These results confirm that tum- variants can elicit a syngeneic rejection response against tumors that are apparently devoid of transplantation immunogenicity. The experimental conditions and the results make it likely but not certain that the tumor-associated antigen detected on leukemia TH was present on the primary tumor.
Mediators from cloned T helper cell lines affect immunoglobulin expression by B cells.
Authors: Paige C J , Schreier M H , Sidman C L
Published: May 2006

When cloned T helper cells encounter antigen presented by I-A-compatible macrophages, soluble mediators are produced that affect the differentiation and activation of normal B lymphocytes and cell lines of the B lineage. Exposure to such T cell culture supernatants causes two effects in the murine 70Z/3 cell line, which represents a pre-B stage of differentiation. These cells begin to synthesize Ig light chains and gain membrane Ig that is detectable by immunofluorescence. Two other effects are seen after similar treatment of the WEHI-279.1 murine cell line, which represents a mature, Ig+ B cell. These cells shift the ratio of mu chains produced from mostly membrane to mostly secretory type and begin to secrete large amounts of IgM, which can be detected either by biosynthetic radiolabeling followed by immunoprecipitation or by a staphylococcal protein A plaque assay. The majority also die. Similar to WEHI-279.1, normal small resting B cells also show the shift from membrane mu to secretory mu and are activated to Ig secretion after exposure to these supernatants. These results show that products from T cell immune reactions exert multiple effects on B cell development and activation, at several stages of teh B cell developmental pathway. The observed change range from nuclear processes, including gene transcription and RNA splicing, to such post-translational aspects as protein processing, catabolism, membrane architecture, and cell survival.
Evolution of influenza A and B viruses: conservation of structural features in the hemagglutinin genes.
Authors: Krystal M , Elliott R M , Benz E W , Young J F , Palese P
Published: May 2006

The complete nucleotide sequence of the hemagglutinin (HA) gene of a type B influenza virus (B/Lee/40) was obtained by using cloned cDNA derived from the RNA segment. The gene is 1,882 nucleotides long and can code for a protein precursor of 584 amino acids. Structural features common to type A virus HAs are also conserved in the B virus HA. These include a hydrophobic signal peptide, hydrophobic NH2 and COOH termini of the HA2 subunit, and a HA1/HA2 cleavage site involving an arginine residue. The sequence of the B HA gene and its deduced amino acid sequence were compared to those of a type A influenza virus (A/PR/8/34). When these two genes were aligned, it was found that 24% of the amino acids in the HA1 subunits and 39% of the amino acids in the HA2 subunits are conserved. This degree of relatedness between type B virus and type A virus HAs (intertypic comparison) is similar to the homologies observed among certain type A virus HAs (intratypic comparison). A close evolutionary relationship is therefore suggested between the HAs of type A and type B influenza viruses.
Isolation and expression of cloned hook protein gene from Caulobacter crescentus.
Authors: Ohta N , Chen L S , Newton A
Published: May 2006

Previous genetic analysis of Caulobacter crescentus showed that the periodic synthesis of hook protein, flagellin A, and flagellin B, the major flagellar subunits, is coupled in some way to chromosome replication. To examine the regulation of flagellar gene expression at the molecular level, we isolated the gene that codes for the 72,000-dalton hook protein. A specific 125I-labeled anti-hook protein IgG was used to screen a hybrid lambdaL47.1 bank of 4,500 clones and to compare peptide maps of the cloned gene product with purified hook protein. Restriction analysis of DNA from the positive lambda clones and plasmid subclones showed that the structural gene for the hook protein is contained on a 2.3-kilobase (kb) BamHI fragment. The direction of transcription was established by demonstrating the inducibility of hook protein gene in strains with the 2.3-kb fragment fused to the Escherichia coli lipoprotein gene-lactose gene promoter-operator region of pIN-II. Preliminary genomic analysis showed that the hook gene occupies a single location on the C. crescentus chromosome. These results suggest that the periodic expression of the hook protein gene in the cell cycle does not involve a major or persistent rearrangement of the 2.3-kb coding sequence during the cell cycle.
Regulation of expression of the Escherichia coli dnaG gene and amplification of the dnaG primase.
Authors: Wold M S , McMacken R
Published: May 2006

We have isolated lambda transducing phages carrying the Escherichia coli primase gene (dnaG) and mapped restriction sites in the cloned bacterial DNA segments. Several different DNA fragments containing the dnaG gene were inserted into multicopy plasmids. An analysis of the primase levels in cells harboring such plasmids indicates that sequences far upstream from the dnaG gene are required for optimal primase expression. Using this knowledge, we constructed a plasmid with a thermoinducible copy-number, pRLM61, which was employed to amplify intracellular primase levels approximately 100-fold. The dnaG gene is transcribed clockwise with respect to the E. coli genetic map, and a HindIII site located 180 base pairs upstream from the dnaG gene separates the gene from its primary promoter. An apparent transcription termination signal is positioned 30-70 base pairs in front of the primase gene. Transcription proceeds past this strong terminator only when RNA polymerase has first transcribed the bacterial DNA segment proximal to the HindIII site. We suggest that primase expression in E. coli is positively regulated by a mechanism of transcription antitermination mediated by a bacterial factor. We propose, furthermore, that the neighboring structural genes for primase and for the sigma subunit of RNA polymerase are coordinately regulated as part of an operon. This arrangement may enable the bacterial cell to readily control the level of initiation of DNA and RNA synthesis and thus to respond quickly and efficiently to changing conditions.
Induction by dimethyl sulfoxide of Fc fragment-mediated phagocytosis in the mouse macrophage-like cell line P388D1.
Authors: Takasaki S , Leive L
Published: May 2006

The murine macrophage-like cell line P388D1 has Fc fragment receptors but cannot carry out Fc receptor-mediated phagocytosis. We now find that it gains this ability when exposed to 1.5% dimethyl sulfoxide for 2 days. Binding by Fc receptors and phagocytosis of latex beads are unaffected. Growth eventually slows, and the activities of two lysosomal enzymes--beta-glucuronidase and acid phosphatase--increase, but these increases occur after the induction of Fc receptor-mediated phagocytosis. Cycloheximide sufficient to inhibit protein synthesis inhibits the effect of dimethyl sulfoxide. All changes are reversed when dimethyl sulfoxide is removed. This system may facilitate study of differentiation processes affecting Fc receptor-mediated phagocytosis.
Chloroplast DNA evolution and phylogenetic relationships in
Lycopersicon
Authors: Palmer Jeffrey D. , Zamir Daniel
Published: May 2006

Chloroplast DNA was purified from 12 accessions that represent most of the species diversity in the genus
Lycopersicon
(family Solanaceae) and from 3 closely related species in the genus
Solanum
. Fragment patterns produced by digestion of these DNAs with 25 different restriction endonucleases were analyzed by agarose gel electrophoresis. In all 15 DNAs, a total of only 39 restriction site mutations were detected among 484 restriction sites surveyed, representing 2,800 base pairs of sequence information. This low rate of base sequence change is paralleled by an extremely low rate of convergent change in restriction sites; only 1 of the 39 mutations appears to have occurred independently in two different lineages. Parsimony analysis of shared mutations has allowed the construction of a maternal phylogeny for the 15 accessions. This phylogeny is generally consistent with relationships based on morphology and crossability but provides more detailed resolution at several places. All accessions within
Lycopersicon
form a coherent group, with two of the three species of
Solanum
as outside reference points. Chloroplast DNA analysis places
S. pennellii
firmly within
Lycopersicon
, confirming recent studies that have removed it from
Solanum
. Red-orange fruit color is shown to be a monophyletic trait in three species of
Lycopersicon
, including the cultivated tomato,
L. esculentum
. Analysis of six accessions within
L. peruvianum
reveals a limited amount of intraspecific polymorphism which, however, encompasses all the variation observed in
L. chilense
and
L. chmielewskii
. It is suggested that these latter two accessions be relegated to positions within the
L. peruvianum
complex.
Monoclonal antibodies to human prostatic acid phosphatase: probes for antigenic study.
Authors: Lillehoj H S , Choe B K , Rose N R
Published: May 2006

Hybrid cell lines producing monoclonal antibodies against human prostatic acid phosphatase [PAPase; orthophosphoric-monoester phosphohydrolase (acid optimum), EC 3.1.3.2) were prepared by the fusion of mouse myeloma cells with the spleen cells of PAPase-immunized BALB/c mice. Approximately 23% of the hybrid cells initially plated after cell fusion produced specific antibodies: 34 microcultures were cloned, and 8 eventually yielded stable cell lines. The monoclonal antibodies produced by these eight hybridomas were characterized for their isotypes, isoelectric points, concentrations, and affinities. All of the eight monoclonal antibodies exhibited strict specificity for PAPase as determined by radioimmunoassay and immunohistochemical methods. These antibodies were used as probes for the antigenic mapping of this enzyme, and three nonoverlapping determinants were recognized. Further binding studies with PAPase fragments, generated by cleavage with a submaxillaris protease, showed that those three determinants are clustered on one fragment of PAPase. These monoclonal antibodies may be useful in refinement of clinical immunoassays of PAPase or immunohistological study of PAPase-synthesizing cells.
Hormone receptor modulates the regulatory component of adenylyl cyclase by reducing its requirement for Mg2+ and enhancing its extent of activation by guanine nucleotides.
Authors: Iyengar R , Birnbaumer L
Published: May 2006

N-Ethylmaleimide treatment of rat liver plasma membranes results in an adenylyl cyclase (EC 4.6.1.1) system that shows no measurable cyclizing activity but retains both an active glucagon receptor and a receptor-sensitive regulatory component N as assessed by reconstitution into cyclase-negative (cyc-) membranes from S49 murine lymphoma. Treatment of such N-ethylmaleimide-treated membranes, termed C- liver membranes, with guanosine 5'-[gamma-thio]triphosphate (GTP[gamma S] ) and Mg2+, followed by the removal of GTP[gamma S] by washing, yields an activated N which upon mixing with cyc- S49 membranes reconstitutes the cyc- S49 membrane adenylyl cyclase in the absence of added GTP[gamma S]. It was found that GTP[gamma S] activation of the N at saturating concentrations of GTP[gamma S] is slow at low Mg2+ concentration and accelerated by increasing Mg2+ concentrations. Addition of glucagon during the activation results in a lowering of the Mg2+ requirement for full activation from 25 mM to around 10 muM and in concomitant increases in both the rate and the extent of N activation. In contrast to its dramatic effect on Mg2+ requirement, glucagon has little (less than 2-fold) effect on the GTP[gamma S] requirement of N activation. These experiments indicate that the glucagon receptor facilitates activation of N by: (i) decreasing the apparent Km of N for Mg2+, and (ii) increasing the extent of activation that can be elicited by saturating concentrations of guanine nucleotide. It is postulated that the mechanism by which Mg2+ and receptors facilitate N activation involves dissociation of n alpha activated ADP-ribosylatable subunits (with guanine nucleotide bound to them) from n beta non-ADP-ribosylatable subunits (with receptor and Mg2+ bound to them).
Sarcomeric myosin heavy chain is coded by a highly conserved multigene family.
Authors: Nguyen H T , Gubits R M , Wydro R M , Nadal-Ginard B
Published: May 2006

pMHC25, a recombinant plasmid containing myosin heavy chain (MHC) cDNA sequences from differentiated myotubes of the L6E9 rat cell line, has been shown to hybridize to all sarcomeric MHC mRNAs so far tested but not to nonsarcomeric MHC mRNAs. In addition, pMHC25 hybridizes to multiple restriction endonuclease-digested fragments of rat genomic DNA corresponding to different MHC genomic sequences. Thus, the MHC gene represented by pMHC25 is a member of a sarcomeric MHC multigene family that has regions of sequence homology shared among its members. This sarcomeric MHC multigene family has been estimated to be composed of a minimum of seven genes, some of which are polymorphic in the rat. We have also determined that pMHC25 hybridizes to MHC gene sequences in genomic DNA of all species that have striated muscle, ranging from nematodes to man. Sarcomeric MHC genes, therefore, have been horizontally and vertically conserved in evolution. Additionally, we have used the pMHC25 plasmid to demonstrate that MHC genes do not undergo rearrangement or amplification during muscle cell differentiation.