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Mol. Hum. Reprod. Advance Access published online on March 27, 2006
Molecular Human Reproduction, doi:10.1093/molehr/gal028
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© The Author 2006. Published by Oxford University Press on behalf of the European Society of Human Reproduction and Embryology. All rights reserved. For Permissions, please email: journals.permissions@oxfordjournals.org
Received January 9, 2006
Revised February 13, 2006
Accepted February 18, 2006

Article

Identification and characterization of oviductal glycoprotein-binding protein partner on gametes: epitopic similarity to non-muscle myosin IIA, MYH 9

K.M. Kadam 1, S.J. D’Souza 1, A.H. Bandivdekar 1, and Usha Natraj 1 *

1 National Institute for Research in Reproductive Health, Indian Council for Medical Research, Mumbai, Maharashtra, India

* To whom correspondence should be addressed.
Usha Natraj, E-mail: ushan3{at}gmail.com


  Abstract

The mammalian estrogen induced oviductal glycoprotein (OGP) has been known to associate with capacitated sperm, oocytes and developing embryos. This study aimed to identify the putative binding partner of OGP on gametes using N-terminal peptide of bonnet monkey (Macaca radiata) OGP, Nmon, as bait. A protein(s) of molecular size ~54 kDa was detected by far-western blot analysis of detergent solubilized human sperm proteins. MALDI-TOF mass spectra analysis of ~54 kDa tryptic peptides gave a significant hit to non-muscle myosin heavy chain. Biochemical characterization of ~54 kDa was done with antibodies specific to non-muscle myosin IIA, MYH9. The ~54 kDa protein, possible breakdown product of MYH9, immuno-reacted with MYH9 antibody in western blot analysis. OGP binding to ~54 kDa could also be demonstrated in far-western blot analysis of detergent solubilized human sperm proteins and nuclear matrix intermediate filament (NM-IF) preparations from human sperm and mouse oocytes. Far-western blot analysis of MYH9 enriched by immunoprecipitation identified the native ~220 kDa protein as OGP-binding partner. The identical and characteristic immunogold localization pattern of Nmon and MYH9 on sperm NM-IF preparation substantiated these findings. The results suggest that OGP binds to both gametes through its interaction with MYH9 through the non-glycosylated N-terminal conserved region of OGP, spanning the residues 11-137.

Keywords: cytoskeleton/NM-IF/MYH9/oviductal glycoprotein.
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