-LOCATION, ISOLATION AND CHARACTERIZATION OF PROTEIN WITH THIOL GROUPS DETECTED DURING MATURATION IN SPERM

CABRILLANA M A; MONCLUS M A; VINCENTI A E; FORNES M W

Potrero de los Funes - San Luís

Congreso; XXIV REUNIÓN CIENTÍFICA ANUAL DE LA SOCIEDAD DE BIOLOGÍA DE CUYO Y IV REUNIÓN CIENTÍFICA DE LA SOCIEDAD ARGENTINA DE MICROSCOPIA (SAMIC); 2006

Sociedad de Biología de Cuyo

Location, isolation and characterization of protein with thiol groups detected during maturation in sperm. Cabrillana ME, Monclus MA, Vincenti AE and Fornés Miguel Walter. IHEM, FCMédicas. UNCuyo - CONICET. Mendoza. mcabrillana@yahoo.com.ar Oocyte fertilization involve maturate and capacitated sperm. The sperm support maturation during the epididymal transit and capacitation during the stadium in the female genital tract.  Changes in thiol oxidation, to disulfide bonds, in sperm proteins are verified during epididymal trip. Instead, the capacitation process is marked by an increment in tyrosine phosphorilated sperm protein (p-Y).  Using sonication and ultracentrifugation in sucrose gradient we fractionated and isolate sperm head and tails from mature vs. immature or capacitated vs. non-capacitated sperm. Them incubation of fractions with monobromobimane (mbb) permit to stain thiol proteins. The cell location of specific proteins was established in sperm head or tail by fluorescence microscopy. The molecular weight and thiol content of protein were determined by standard SDS-Page. Parallel gel was run to perform and immune-blot for p-Y. Some samples were incubated with detergents (1% Triton X100) and soluble vs non-soluble proteins were also analyzed with the protocol above mention. Phosphorilated and mbb marked proteins were coincident in mature and capacitated sperm and were isolated in soluble fraction of sperm. Results indicate that same proteins are stabilized during maturation by thiol changes and phosphorilated during capacitation.