Primate recombinant zona pellucida proteins expressed in Escherichia coli bind to spermatozoa

Gahlay, Gagandeep K. ; Srivastava, Neelu ; Govind, Chhabi K. ; Gupta, Satish K. (2002) Primate recombinant zona pellucida proteins expressed in Escherichia coli bind to spermatozoa Journal of Reproductive Immunology, 53 (1-2). pp. 67-77. ISSN 0165-0378

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To delineate the role of individual zona pellucida (ZP) glycoproteins during sperm–oocyte interaction, bonnet monkey (bm; Macaca radiata) ZPA (bmZPA), ZPB (bmZPB), and ZPC (bmZPC) have been cloned without native signal sequence and transmembrane-like domain, and expressed in Escherichia coli. Recombinant proteins have been purified from the inclusion bodies in presence of low concentration of chaotropic agent (2 M urea) and high pH (pH 12), and subsequently refolded in presence of oxidized and reduced glutathione. Binding of the recombinant refolded zona proteins to bonnet monkey spermatozoa in an indirect immunofluorescence assay revealed that recombinant bmZPC binds to the head region of the capacitated spermatozoa but does not bind to the acrosome reacted spermatozoa. Recombinant bmZPB binds to the principal segment of the acrosomal cap of capacitated bonnet monkey spermatozoa. After induction of acrosome reaction by calcium ionophore A23187, the binding of recombinant bmZPB shifts to the equator, post-acrosome and midpiece of the spermatozoa. bmZPA binds to the principal segment of capacitated spermatozoa but the binding shifts to the equatorial segment, tip of the inner acrosomal membrane and midpiece in acrosome reacted spermatozoa. These studies suggest that polypeptide backbone is sufficient for the binding of ZPA, ZPB and ZPC to spermatozoa in non-human primates. Further studies with recombinant glycosylated zona proteins will help in delineating the role of carbohydrate moieties for higher affinity binding of the ligand to spermatozoa and subsequent signal transduction pathways.

Item Type:Article
Source:Copyright of this article belongs to Elsevier Science.
Keywords:Recombinant Zona Pellucida Glycoproteins; E-coli Expression; Sperm Binding
ID Code:72139
Deposited On:28 Nov 2011 06:15
Last Modified:28 Nov 2011 06:15

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