Characterization of a 5700 - dalton sperm motility inhibitor from seminal plasma of boar

Abstract

This chapter discusses Saccharomyces cerevisiae proteins that promote hybrid DNA formation in vitro. In order to identify additional factors that are involved in hybrid DNA formation in Saccharomyces cerevisiae an in vitro stimulation of strand-exchange protein 1 (SEPl) is used to identify proteins that reconstitute strand-exchange activity in reactions containing limiting amounts of SEPl. Two proteins are identified that functionally interact with SEPl. First, an Mr 34,000 single-stranded, DNA-binding protein stimulated the reaction by lowering the requirement for SEPl about three- to four-fold. This protein is a fragment of the large subunit of a heterotrimeric complex called yRPA (yRFA), which is thought to be the functional eukaryotic equivalent of prokaryotic single-stranded, DNA-binding proteins. Additional analysis has demonstrated that the intact three-subunit form of yeast RPA will also stimulate SEPl, similar to the Mr 34,000 fragment of the large subunit of RPA. The gene encoding this protein (RPA1) is essential for growth. Second, an Mr 33,000 polypeptide, termed Stimulatory Factor 1 (SF1), dramatically stimulated the SEPl-catalyzed reaction by lowering the requirement for SEPl about 300-fold. This protein specifically stimulated SEPl in a way that is quantitatively and qualitatively different from that observed with other strand-exchange stimulatory proteins.