Functional activity of human ZP3 primary sperm receptor resides toward its C-terminus

Bansal, Pankaj ; Chakrabarti, Kausiki ; Gupta, Satish K. (2009) Functional activity of human ZP3 primary sperm receptor resides toward its C-terminus Biology of Reproduction, 81 (1). pp. 7-15. ISSN 0006-3363

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Zona pellucida glycoprotein 3 (ZP3) has been ascribed as a putative primary sperm receptor during fertilization in humans. Herein, attempts have been made to delineate the functional domain of human ZP3. ZP3 has been cloned and expressed in a baculovirus expression system as Nterminal fragments (amino acid [aa] residues 1-175 [pAc-ZP3(1-175 aa)] and 23-175 [pBg-ZP3(23-175 aa)]) and as C-terminal fragments (aa residues 214-305 [pBg-ZP3(214-305 aa)] and 214-348 [pBg-ZP3(214-348 aa)]). ZP3 encompassing both N- and C-terminal fragments corresponding to aa residues 1-370 (pAc-ZP3[1-370 aa]) has also been expressed. Lectin-binding analysis with these recombinant proteins revealed the presence of N- and O-linked glycosylation. Significant induction of acrosomal exocytosis was observed when capacitated sperm were incubated with pBg-ZP3(214-348 aa), pBg-ZP3(214-305 aa), and pAc-ZP3(1-370 aa) (P < 0.05), whereas incubation with pAc-ZP3(1-175 aa) and pBg-ZP3(23-175 aa) failed to do so under similar experimental conditions. However, N- and C-terminal fragments labeled with fluorescein isothiocyanate revealed binding to the anterior head of capacitated human spermatozoa. Escherichia coli-expressed ZP3 C-terminal fragments and chemically deglycosylated pBg-ZP3(214-348 aa) failed to induce a significant (P < 0.05) increase in acrosomal exocytosis, suggesting the relevance of glycosylation in imparting functional activity to ZP3 C-terminal fragments. pBg-ZP3(214-348 aa)-mediated induction of acrosomal exocytosis is regulated by Gi protein, extracellular calcium, GABA(A) [gamma aminobutyric acid (A)] receptor-mediated Cl- channel, and T-type voltage-operated calcium channels. Taken together, the results of these studies suggest that the functional activity of human ZP3 resides in its C-terminal domain.

Item Type:Article
Source:Copyright of this article belongs to Society for the Study of Reproduction.
Keywords:Acrosome Reaction; Baculovirus Expression System; Recombinant Human ZP3 Fragments: Sperm; Zona Pellucida Domain
ID Code:12037
Deposited On:10 Nov 2010 04:50
Last Modified:16 May 2016 21:26

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