ANALYSIS OF POLYSULFATE-BINDING DOMAINS IN PORCINE PROACROSIN, A PUTATIVE ZONA ADHESION PROTEIN FROM MAMMALIAN SPERMATOZOA

Proacrosin is one of the major proteins found within the acrosomal ves icle of mammalian spermatozoa. Previous work has shown that it binds n on-enzymically and with high affinity to polysulfate groups on zona pe llucida glycoproteins (ZPGPs) thereby leading to the hypothesis that a t fertilization it functions as a secondary ligand molecule to retain acrosome-reacted spermatozoa on the surface of the egg. In the present work we have investigated the nature and extent of the polysulfate bi nding domain on boar sperm proacrosin using a combination of group-spe cific modifying reagents, fragmentation analysis, peptide synthesis an d expression of deletion recombinants in E. coli bacteria. Taken overa ll, our results show that arginine, lysine and histidine residues loca ted between Gly 93 and Ala 275, together with the participation of His 47 and Arg 50, are necessary for maximum polysulfate binding activity . The secondary and tertiary structure of this central peptide domain is also important to ensure correct alignment of basic residues with c omplementary sulfate groups on ZPGPs. Proacrosin, therefore, has many properties in common with other polysulfate binding proteins, such as antithrombin III and sea urchin sperm bindin, in having a canformation -dependent domain containing basic amino acids that mediates specific protein-protein interactions. These observations strengthen the hypoth esis that proacrosin is a multifunctional protein with a major role as a ligand molecule at fertilization.