p95, the major phosphotyrosine-containing protein in mouse spermatozoa, is a hexokinase with unique properties.
Mouse sperm contain a major phosphotyrosine-containing protein of M(r) 95,000 (nonreducing conditions) which has been implicated as a sperm membrane receptor for the egg zona pellucida glycoprotein, ZP3 (Leyton, L., and Saling, P. (1989) Cell 57, 1123-1130; Leyton, L., LeGuen, P., Bunch, D., and Saling, P. (1992) Proc. Natl. Acad. Sci. U. S. A. 89, 11692-11695). This protein was purified and subjected to limited tryptic digestion and subsequent amino acid analysis. Three sequenced peptides revealed 100% amino acid identity to a mouse hepatoma hexokinase (Arora, K. K., Fanciulli, M., and Pederson, P. L. (1990) J. Biol. Chem. 265, 6481-6488). The purified protein, which migrated at M(r) 116,000 under reducing conditions (p95/116), reacted with an antiserum to the purified rat brain hexokinase, type 1, and comigrated on sodium dodecyl sulfate-polyacrylamide gel electrophoresis with the purified rat brain enzyme under both nonreducing and reducing conditions. Unlike p95/116, the rat brain enzyme was not a phosphotyrosine-containing protein. The p95/116 protein could be immunoprecipitated with the hexokinase antiserum or an O-phosphotyrosine antibody. Limited tryptic digestion of the purified p95/116 and the rat brain enzyme generated subsets of identical peptides which reacted with the hexokinase antiserum. However, p95/116 also contained phosphotyrosine-containing peptides that were not present in the rat brain hexokinase. When different mouse tissues were probed with the hexokinase antiserum all tissues, with the exception of liver, contained immunoreactive protein. In contrast, only sperm and testis possessed a phosphotyrosine-containing form of hexokinase. These data suggest that the germ cell component of the testis possesses a unique tyrosine-phosphorylated form of hexokinase.
Kalab
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Department of Obstetrics and Gynecology, University of Pennsylvania School of Medicine, Philadelphia 19104-6080.
Visconti
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Leclerc
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Kopf
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D000595:Amino Acid Sequence; D000818:Animals; D015153:Blotting, Western; D001921:Brain / Q000201:enzymology; D002462:Cell Membrane / Q000201:enzymology; D004591:Electrophoresis, Polyacrylamide Gel; D006593:Hexokinase / Q000737:chemistry* / Q000302:isolation & purification; D008297:Male; D051379:Mice; D008815:Mice, Inbred Strains; D008969:Molecular Sequence Data; D008970:Molecular Weight; D009928:Organ Specificity; D010446:Peptide Fragments / Q000737:chemistry; D010750:Phosphoproteins / Q000737:chemistry* / Q000302:isolation & purification; D019000:Phosphotyrosine; D051381:Rats; D013094:Spermatozoa / Q000201:enzymology*; D013737:Testis / Q000201:enzymology; D014443:Tyrosine / Q000031:analogs & derivatives* / Q000032:analysis
D010446:Peptide Fragments; D010750:Phosphoproteins; D019000:Phosphotyrosine; D014443:Tyrosine; D006593:Hexokinase