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Biomedical subjects

J H Shaper

Publications and source records attributed to J H Shaper.

At least 91 records · Page 5Linked to original sources
PubMed1981

Physical and chemical properties of the major envelope glycoprotein gp85 from avian myeloblastosis virus.

The major envelope glycoprotein gp85 of avian myeloblastosis virus, observed by electron microscopy as nearly spherical knobs projecting from the virus surface, was purified to homogeneity by gel filtration in 6 M guanidinium chloride followed by ion-exchange chromatography. The purified glycoprotein has a molecular weight of 80 000 from sedimentation equilibrium analysis. Glycoprotein gp85 contains approx. 45% carbohydrate including 25% N-acetylglucosamine, while the remaining weight consists of a polypeptide chain of approx. 45 000 daltons. Based on the oligosaccharide chain molecular weight data of Lai and Duesberg (Lai, M.M.C. and Duesberg, P.H. (1972) Virology 50, 359-372), the carbohydrate is calculated to be distributed between seven to nine oligosaccharide side chains. No self-association of gp85 was observed up to 2.0 mg/ml in dilute salt solution. The hydrodynamic properties of gp85 in dilute salt solution indicate a highly elongated molecule with an axial ratio of 7. One structural model which reconciles the hydrodynamic properties of gp85 with the nearly spherical architecture observed by electron microscopy requires the organization of the polypeptide chain and approx. 50% of the carbohydrate into a globular form. The remaining covalently linked oligosaccharides would by necessity extend outwardly from the globular structure as randomly oriented chains.

Amino Acids↗
PubMed1980

Serum UDP-galactosyl transferase as a potential biomarker for breast carcinoma.

UDP-galactose:N-acetylglucosamine galactosyltransferase (GT) is a membrane-bound enzyme active in the biosynthesis of the carbohydrate moiety of glycoproteins and glycolipids. A soluble form of GT, present in human serum, has recently been found to be elevated in the presence of various neoplasms. In this study, GT levels were measured in randomized serum samples obtained from normal controls (group I, n = 49), patients with benign breast disease (group II, n = 46), disease controls (group III, n = 50), patients with primary breast carcinoma (group IV, n = 53), and untreated metastatic breast cancer (group V, n = 23). Although substantial serum GT elevations were observed in individual control patients with active inflammatory or metabolic diseases, the mean GT levels were significantly higher in the groups with breast carcinoma (P < 0.001, 0.001, 0.02; P < 0.001, 0.001, 0.001 for groups IV and V vs groups, 1, II, and III, respectively). Furthermore, when serum GT levels were correlated with the properative clinical stage of breast cancer, significant elevations were found in 14.3% (3/21) of stage I, 66.7% (8/12) of stage II, 78.6% (11/14) of stage III, and 96.5% (28/29) of stage IV patients. These data indicate that serum GT levels are elevated in the presence of breast carcinoma and that the enzyme elevations correlate positively with the clinical stage of disease. Serum GT may be potentially useful in the detection of recurrent breast carcinoma and as a marker of tumor response to therapy for advanced disease.

Breast Neoplasms↗
PubMed1978

Separation of human renal renin and pseudorenin by affinity chromatography on hemoglobin-Sepharose-2B.

Human renal renin (EC 3.4.99.19) and pseudorenin were easily separated in a single step by affinity chromatography on hemoglobin-Sepharose-2B. Renin and pseudorenin were monitored by their actions on crude and partially purified hog protein renin substrates at neutral and acidic pH and on synthetic labelled polymeric renin substrate. Under the conditions employed (0.1 M sodium acetate (pH 3.5)/1 M sodium chloride at 4 degrees C) renin does not bind to the affinity adsorbent while pseudorenin is effectively bound and can be eluted only after raising the pH to 6.5. Pseudorenin-free renin prepared by this method is devoid of proteolytic activity toward hemoglobin. The chromatographic behaviour of renal pseudorenin on hemoglobin-Sepharose-2B is similar to that of cathepsin D.

Chromatography, Affinity↗
PubMed1977

Accessibility of the carbohydrate moiety of membrane-boound rhodopsin to enzymatic and chemical modification.

Galactose was specifically inserted into the carbohydrate moiety of rhodopsin by incubating retinal disk membranes with UDP-galactose: N-acetylglucosamine galactosyltransferase. The stoichiometry of labeling ranged from 1.2 to 1.8 (average = 1.5) residues of galactose per molecule of rhodopsin, indicating that some or all of the oligosaccharide chains of membrane-bound rhodopsin are readily accessible to enzymatic modification. These modified membranes were treated with galactose oxidase to generate an aldehyde at the C-6 position of the inserted galactose units. The enzymatically-oxidized membranes were then reacted with dansyl hydrazide to yield a fluorescent hydrazone which is sufficiently stable to permit spectroscopic analysis. This procedure for the specific attachment of a spectroscopic probe should be applicable to a wide variety of membrane glycoproteins.

Animals↗
PubMed1977

Classification of Bacillus subtilis flagellins.

Purified flagellins derived from 16 strains of Bacillus subtilis were classified into at least five distinct groups on the basis of their reaction with antiflagellar filament antibody and antiflagellin antibody. This classification was in good accord with that derived independently on the basis of amino acid analyses of the flagellins. Flagellar antigenicity appears to provide a useful typological character in classifying B. subtilis strains.

Amino Acids↗
PubMed1976

Amino acid sequence of flagellin of Bacillus subtilis 168. I. Cyanogen bromide peptides.

Sequence studies are presented on the cyanogen bromide peptides derived from the flagellin of Bacillus subtilis 168. Eight unique CNBr peptides, ranging in length from 4 to 113 residues, were isolated in pure state. These peptides accounted for the amino acid composition of flagellin. The NH2-terminal methionyl residue of the protein reacted only partially with CNBr. Partial cleavage was observed at a Met-Glu bond (residues 22 to 23) of the protein.

Amino Acid Sequence↗
PubMed1976

Amino acid sequence of flagellin of Bacillus subtilis 168. II. Tryptic peptides from maleylated flagellin.

Fifteen pure peptides, ranging in length from 2 to 141 residues, were isolated from the tryptic digest of maleylated flagellin. Studies of these peptides in conjunction with the data on the eight cyanogen bromide peptides described earlier (Chang, J. Y., DeLange, R. J., Shaper, J. H., and Glazer, A. N. (1976) J. Biol. Chem. 251, 695-700) established the order of three of the CNBr peptides at the NH2 terminus and of three at the COOH terminus of flagellin.

Amino Acid Sequence↗
PubMed1976

Amino acid sequence of flagellin of Bacillus subtilis 168. III. Tryptic peptides, N-bromosuccinimide peptides, and the complete amino acid sequence.

Of the 28 expected tryptic peptides from Bacillus subtilis 168 flagellin, 24 were isolated and sequenced. Several overlapping tryptic peptides were also characterized. Studies were also performed on two peptides of 142 and 162 residues isolated after cleavage of the flagellin molecule at the single tyrosine residue (residue 142) with N-bromosuccinimide. These studies together with the previous data on the cyanogen bromide peptides and the tryptic peptides from maleylated flagellin permitted the complete amino acid sequence to be established: (see article). The primary structure reveals no obvious regularities or major repetitions of homologous sequences. Hydrophobic residues are distributed randomly in the amino acid sequence. However, the distribution of charged residues is strikingly asymmetric. The NH2-terminal region (residues 1 to 101) possesses a net charge of 6 plus, the middle of the molecule (residues 102 to 203), a net charge of 9 minus, and the COOH terminal region (residues 204 to 304), a net charge of 4 minus.

Amino Acid Sequence↗
PubMed1975

Cross-linking of the components of lactose synthetase with dimethylpimelimidate.

The cross-linking of the two components of lactose synthetase, alpha-lactalbumin and a galactosyltransferase, with dimethylpimelimidate was examined. The extent of the cross-linking at pH 8.1 was found to be dependent upon the presence of substrates or inhibitors for the galactosyltransferase. N-acetylglucosamine and mixtures of either N-acetylglucosamine, Mn-2+ and UDP, or UDP-galactose and Mn-2+ promoted the formation of cross-linked species. Glucose or a mixture of UDP and Mn-2+ were much less effective in promoting cross-linking. Two types of intermolecularly cross-linked species of alpha-lactalbumin and the galactosyltransferase were obtained. Each was a 1:1 cross-linked complex of alpha-lactalbumin and either of the two forms of the transferase with molecular weights of about 42,000 and 48,000, respectively. Cross-linked complexes were not observed with more than 1 molecule each of alpha-lactalbumin and the transferase. The cross-linked complexes were obtained in homogeneous form by gel filtration on Sephadex and absorption of uncross-linked enzyme by affinity chromatography on alpha-lactalbumin-Sepharose in the presence of N-acetylglucosamine. They migrated on gel electrophoresis in sodium dodecyl sulfate with mobilities in accord with their predicted molecular weights as 1:1 complexes of alpha-lactalbumin and the transferase. The amino acid composition of the cross-linked complex was in reasonable agreement with the expected composition of a 1:1 mixture of alpha-lactalbumin and galactosyltransferase. The enzymic properties of the cross-linked and uncross-linked enzymes were compared. The cross-linked complex had a much higher intrinsic lactose synthetase activity than did uncross-linked enzyme although only about 1% of the potential activity of uncross-linked enzyme in the presence of optimal concentrations of alpha-lactalbumin. The lactose synthetase activity of the cross-linked complex, however, was unaffected by exogenous alpha-lactalbumin. In addition, the complex readily catalyzed the transfer of galactose from UDP-galactose to xylose in the absence of exogenous alpha-lactalbumin. The N-acetyllactosamine synthetase activity of the complex was low compared to its activity with other monosaccharides. Ovalbumin, which is a good acceptor for the uncross-linked transferase, was not an acceptor for the cross-linked complex. Kinetic studies of the complex suggest that its modified catalytic activity is not the result of the modification by dimethylpimelimidate but reflects the expected effects of is provided, and that

Amino Acids↗