Isolation and characterization of the sperm surface proteins and their binding studies with zona pellucida glycoproteins

0380453 - BTÚ 2013 CZ eng A - Abstract
Zigo, Michal - Jonáková, Věra - Postlerová, Pavla
Isolation and characterization of the sperm surface proteins and their binding studies with zona pellucida glycoproteins.
XVII. symposium českých reprodukčních imunologů s mezinárodní účastí. Praha: Biotechnologický ústav v. v. i. AVČR, 2011 - (Pěknicová, J.). s. 65-66
[XVII. symposium českých reprodukčních imunologů s mezinárodní účastí. 26.05.2011-29.05.2011, Žďár nad Sázavou]
R&D Projects: GA ČR(CZ) GA523/09/1793; GA ČR(CZ) GA523/08/H064; GA MŠMT(CZ) 1M06011; GA MZd(CZ) NS10009
Institutional research plan: CEZ:AV0Z50520701
Keywords : monoclonal antibody * zona pellucida * glycoproteins * sperm surface proteins
Subject RIV: CE - Biochemistry

Recognition and the binding of spermatozoa to the zona pellucida is a crucial step in the fertilization process. The sperm-ZP attachment is mediated by complementary molecules on the surface of both gametes and involves the interaction of sperm protein receptors with the ZP structures. Despite longstanding research efforts to identify the sperm proteins that recognize ZP receptors, their precise determination still remains questionable in various mammalian species. Additionally, the exact methods for isolation of sperm membrane proteins have not been properly standardized yet. It has been shown that many proteins receptors are involved in sperm-ZP interaction. Moreover, absence of one specific receptor on the sperm does not have to result in an instant loss of ability to bind with oocyte. Several potential primary sperm receptors for ZP glycoproteins have been investigated in various mammals. The majority of proteins with identified sperm-ZP binding activity belong to the plasma membrane proteins. Sperm surface proteins of ejaculated and capacitated boar sperms were isolated and characterized by 1D/2D protein profiles, glycoprotein staining and substrate zymographic methods for hyaluronidase and proteolytic activity. Interactions between sperm surface proteins of both ejaculated and capacitated sperms with ZP glycoproteins were studied. Results have shown the quantitative and qualitative differences in 1D protein profiles between ejaculated and capacitated sperms, Glycoprotein profiles were roughly the same as 1D protein profiles. Nor hyaluronidase neither proteolytic activity was observed. Binding assay proved that interactions occur between both ejaculated and capacitated sperms. Panel of antibodies against sperm surface proteins was prepared, and used for immunofluorescence test and far western blotting detection. It was observed that some antibodies were binding with intensive signal in fluorescent test, while the signal in far western blotting was rather weak.
Permanent Link: http://hdl.handle.net/11104/0211163