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Titolo:
IMMUNOCYTOCHEMICAL LOCALIZATION AND BIOCHEMICAL-CHARACTERIZATION OF ANOVEL PLASMA MEMBRANE-ASSOCIATED, NEUTRAL PH OPTIMUM ALPHA-L-FUCOSIDASE FROM RAT TESTIS AND EPIDIDYMAL SPERMATOZOA
Autore:
AVILES M; ABASCAL I; MARTINEZMENARGUEZ JA; CASTELLS MT; SKALABAN SR; BALLESTA J; ALHADEFF JA;
Indirizzi:
LEHIGH UNIV,DEPT CHEM,DIV BIOCHEM SCI,111 RES DR BETHLEHEM PA 18015 LEHIGH UNIV,DEPT CHEM,DIV BIOCHEM SCI BETHLEHEM PA 18015 UNIV MURCIA,SCH MED,DEPT CELL BIOL,SECT HISTOL & GEN EMBRYOL E-30071 MURCIA SPAIN
Titolo Testata:
Biochemical journal
, volume: 318, anno: 1996,
parte:, 3
pagine: 821 - 831
SICI:
0264-6021(1996)318:<821:ILABOA>2.0.ZU;2-S
Fonte:
ISI
Lingua:
ENG
Soggetto:
HUMAN ZONA-PELLUCIDA; BINDING PROTEINS; PURIFICATION; CARBOHYDRATE; MANNOSIDASE; FUCOIDIN; SURFACE; HAMSTER; NEOGLYCOPROTEIN; FERTILIZATION;
Tipo documento:
Article
Natura:
Periodico
Settore Disciplinare:
Science Citation Index Expanded
Citazioni:
42
Recensione:
Indirizzi per estratti:
Citazione:
M. Aviles et al., "IMMUNOCYTOCHEMICAL LOCALIZATION AND BIOCHEMICAL-CHARACTERIZATION OF ANOVEL PLASMA MEMBRANE-ASSOCIATED, NEUTRAL PH OPTIMUM ALPHA-L-FUCOSIDASE FROM RAT TESTIS AND EPIDIDYMAL SPERMATOZOA", Biochemical journal, 318, 1996, pp. 821-831

Abstract

1. Immunocytochemical and biochemical techniques have been used to localize and characterize a novel plasma membrane-associated, neutral-pH-optimum alpha-L-fucosidase from rat spermatozoa. Light and electron microscopy specifically localized the fucosidase on the plasma membraneof the convex region of the principal segment of testicular and caudaepididymal sperm heads. Immunoreactivity for alpha-L-fucosidase was also detected in the Golgi apparatus of spermatocytes and spermatids but no immunoreactivity was observed in the acrosome. 2. Fractionation of epididymal sperm homogenates indicated that over 90% of the alpha-L-fucosidase activity was associated with the 48 000 g pellet. This pellet-associated activity could be solubilized with 0.5 M NaCl but not with 0.5% Triton X-100, suggesting that fucosidase is peripherally associated with membranes. Sucrose-density-gradient centrifugation of spermhomogenates indicated that fucosidase was enriched in the plasma membrane-enriched fraction. Analysis of alpha-L-fucosidase on intact epididymal sperm indicated that the enzyme was active, displayed linear kinetics and had a pH-activity curve (with an optimum near 7) which was comparable to that of fucosidase from epididymal sperm extracts. These results further suggest that fucosidase is associated with plasma membranes, and that its active site is accessible to fucoconjugates. Evidence that most of the fucosidase is associated with the exterior of theplasma membrane came from studies in which intact sperm had fucosidase activity comparable to that of sperm sonicates, and from studies in which approx. 90% of the fucosidase activity on intact sperm could be released from the sperm by gentle shaking with 0.5 M NaCl. Isoelectricfocusing indicated that the NaCl-solubilized epididymal sperm fucosidase appears to have one major and one miser isoform with pIs near 7.2 and 5.2, respectively. SDS/PAGE and Western blotting indicated that the NaCl-solubilized extract of epididymal sperm contains two protein bands of 54 and 50 kDa which were highly immunoreactive with the IgG fraction of anti-fucosidase antibodies. Although the function of the novel sperm fucosidase is not known, its specific localization to the plasma membrane of the region of the rat sperm head involved in sperm-egg binding and its high enzymic activity at neutral pH on intact sperm suggest that this enzyme may have a role in sperm-egg interactions.

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Documento generato il 09/07/20 alle ore 12:57:15