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Studies of a sperm antigen recognized by two monoclonal antibodies

University of British Columbia

The monoclonal antibodies HSA-5 and HSA-6 were generated by others in Dr. Lee’s lab against ionophore-treated acrosome reacted human sperm in 1989. They were submitted to the second WHO workshop for interlaboratory evaluations of their potential to be immunocontraceptive vaccine candidates, and were concluded to be Thigh priority candidates in 1989. The two antibodies recognize identical bands on a western blot of crude human sperm extract, which indicate that they recognize the same protein (HSAg-5/6). Indirect immunofluorescent assays performed in our lab have shown that the HSAg-5/6 is locted in the equatorial region of methanol-fixed human sperm, as well as on the head and tail of mouse sperm. The antigen was immunolocalized on human sperm by Homyk et al (1993) who showed that HSAg-5/6 was located on both the inner and outer acrosomal membranes. The corresponding antigen from mouse sperm which binds to HSA-5 and HSA-6 was found to have characteristics identical to those of the human protein as analyzed by western blot assay. The purpose of my project was to study the molecular nature of human HSAg-5/6, and isolate cDNA clones of the corresponding antigen from mouse. This characterization included studying the purified human antigen, and molecular cloning of the homologous protein from a mouse testis cDNA library. From work performed by Dr. T. Yoshiki, it was established that purified HSAg-5!6 is a protein with a molecular mass of approximately 6OkD. My studies established that the isolated cDNA clone expresses the C-terminal region of the protein and accounts for approximately 13% of the total molecular weight, based upon the approximated molecular weight of the cloned fusion protein compared to the molecular weight of the native HSAg 5/6. However, antigenic sites for both monoclonal antibodies lie in this tail end, which suggests that the C-terminus of HSAg-5/6 is exposed to the environment such that an immune response occurs. The studies of HSAg-516 described in this thesis not only supplement our current knowledge regarding sperm-specific proteins, but also aid in development of an immunocontraceptive vaccine.

Thesis/Dissertation

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2009-03-04

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http://hdl.handle.net/2429/5455

Reproductive and Developmental Sciences

University of British Columbia

UBCV

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