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M F Mescher

Publications and source records attributed to M F Mescher.

98 records · Page 6Linked to original sources
PubMed1977

The shape-maintaining component of halobacterium salinarium: a cell surface glycoprotein.

Halobacterium salinarium lacks the rigid peptidogylcan layer necessary for shape-maintenance in most bacteria. Instead, the major cell surface component is a high-molecular-weight glycoprotein resembling those found on the surface of eukaryotic cells. The glycoprotein is extremely acidic and has carbohydrate units attached via both N- and O-glycosidis linkages. Evidence has been obtained that demonstrates that the glycoprotein forms a rigid structural matrix at the cell surface and is responsible for maintenance of the characteristic rod-shaped morphology of these organisms.

Bacitracin↗
PubMed1976

Formation of lipid-linked sugar compounds in Halobacterium salinarium. Presumed intermediates in glycoprotein synthesis.

The ability of bacitracin to inhibit the growth of Halobacterium salinarium suggested that glycosylation of the major envelope component, a high molecular weight glycoprotein, might occur via a pathway involving lipid intermediates. This report demonstrates that the cells have enzymatic activities for formation of lipid-linked sugar compounds having the expected properties of such intermediates. Whole cell homogenate catalyzed the transfer of sugar from UDP-glucose, GDP-mannose, and UDP-N-acetyglucosamine to endogenous lipid acceptors. Two lipid products were formed from UDP-glucose, two from GDP-mannose, and one from UDP-N-acetylglucosamine. Characterization of the partially purified lipids by ion exchange chromatography, thin layer chromatography, and mild acid and base hydrolysis showed the major product in each case to have the properties expected for polyisoprenyl phosphoglucose, polyisoprenyl phosphomannose, and polyisoprenyl pyrophospho-N-acetylglucosamine. Estimates of chain length by thin layer chromatography indicate that the lipid has 11 to 12 isoprene identity as a C55-60-polyisoprenyl pyrophospho-N-acetylglucosamine. The N-acetylglucosamine transferase, present in cell envelope preparations, was partially characterized. The enzyme was found to be extremely halophilic, specifically requiring a high concentration of KCl. Optimum activity was obtained at 4 m KCl and partial substitution of K+ by Na+ resulted in a decrease in activity.

Cell Membrane↗
PubMed1976

Purification and characterization of a prokaryotic glycoprotein from the cell envelope of Halobacterium salinarium.

The glycoprotein which accounts for approximately 50% of the protein and all of the nonlipid carbohydrate of the cell envelope of Halobacterium salinarium (Mescher, M. F., Strominger, J. L., and Watson S. W. (1974) J. Bacteriol. 120, 945-954) has been purified and partially characterized. The glycoprotein has an apparent molecular weight of 200,000, is extremely acidic, and has a carbohydrate content of approximately 10 to 12%. The carbohydrate included neutral hexoses, amino sugar, and uronic acid. Information regarding the number, composition, and mode of attachment of the carbohydrate chains was obtained by isolation and examination of the glycopeptides derived from degradation of cell envelope protein with trypsin and pronase. Trypsin digestion resulted in two glycopeptides. One of these was large (approximately 55,000 daltons) and had most of the neutral hexose linked to it. The carbohydrate moieties consisted of di- and trisaccharides of glucosylgalactose and (uronic acid, glucose)-galactose attached via O-glycosidic linkages between galactose and threonine. The other tryptic glycopeptide had a relatively large heterosaccharide attached to it via an alkaline-stable linkage. The heterosaccharide contained 1 glucose, 8 to 9 galactose, 1 mannose, and 10 to 11 glucosamine residues, and approximately 6 residues of an unidentified amino augar. The alkaline stability of the linkage and the amino acid composition of glycopeptides resulting from Pronase digestion of the tryptic glycopeptide showed that the heterosaccharide was attached to an asparagine residue, presumably via an N-glycosylamine bond to the amide group. The intact glycoprotein has a single N-linked heterosaccharide, 22 to 24 O-linked disaccharides, and 12 to 14 O-linked trisaccharides per molecule. N- and O-glycosidic linkages are the most common carbohydrate-protein linkages in mammalian glycoproteins but, to our knowledge, this is the first report of either type of linkage in a prokaryotic cell envelope protein.

Amino Acids↗
PubMed1976

Structural (shape-maintaining) role of the cell surface glycoprotein of Halobacterium salinarium.

The obligate halophile, Halobacterium salinarium, maintains a rod-shaped morphology under normal growth conditions. Lactoperoxidase(EC 1.11.1.7;donor:hydrogen-peroxide oxidoreductase)-catalyzed iodination and treatment with proteolytic enzymes were used to demonstrate that the recently described envelope glycoprotein (Mescher, M.F. & Strominger, J.L. (1976) J. Biol. Chem. 251, 2005-2014) is the only major cell surface component of this organism. The morphological changes that accompany alteration of the structure of the glycoprotein by growth in the presence of bacitracin or its removal with proteolytic enzymes strongly suggest that it forms a rigid matrix at the cell surface and is responsible for maintenance of the characteristic rod shape.

Bacitracin↗
PubMed1974

Protein and carbohydrate composition of the cell envelope of Halobacterium salinarium.

The isolated cell envelope of Halobacterium salinarium strain 1 contained 15 to 20 proteins that were resolved by polyacrylamide gel electrophoresis in the presence of sodium dodecyl sulfate. All but one of these proteins had molecular weights of 130,000 or less and together accounted for 50 to 60% of the total envelope protein. The remaining 40 to 50% of the envelope protein was accounted for by a single protein with an apparent molecular weight of approximately 194,000 that stained for carbohydrate with periodate-Schiff reagent. The proteolytic enzymes trypsin and Pronase were used to show that the carbohydrate is covalently bound to the protein. Separation of amino sugar- and hexose-containing tryptic peptides by gel filtration indicated that all of the nonlipid carbohydrate of the cell envelope is covalently bound to protein. The results of partial purification by phenol extraction indicated that both the amino sugar and hexose are bound to the 194,000-molecular-weight protein. Exposure of isolated cell envelopes to low salt concentration resulted in solubilization of a majority of the envelope proteins. A relatively small number of proteins, including the high-molecular-weight, carbohydrate-containing protein, remained bound to the sedimentable cell membrane fraction.

Bacterial Proteins↗
PubMedDate not supplied

Cytotoxic T lymphocyte mediated lysis without release of serine esterase.

Cloned cytotoxic T lymphocytes (CTL) can lyse target cells in a Ca2+-dependent or Ca2+-independent fashion, depending on the target cell used. We have used these Ca2+-free assay conditions to determine whether there is serine esterase release in the absence of extracellular Ca2+. We find that under conditions where there is significant target cell killing in the absence of Ca2+, there is no detectable serine esterase release. To the extent that serine esterase and perforin-like molecules reside in the same granules, these results also suggest that target cell killing may occur in the absence of degranulation. Our results provide an example of target cell killing without serine esterase release, and the first indication that lysis and degranulation can be functionally dissociated.

Animals↗