A Basic 18-Amino Acid Peptide Contains the Polysulfate-Binding Domain Responsible for Activation of the Boar Proacrosin/Acrosin System1

Abstract

Proacrosin is the zymogen of acrosin, a serine protease localized in the acrosomal matrix of mammalian sperm. Proacrosin/acrosin binds to solubilized zona pellucida glycoproteins (ZPGs) and various polysulfates in a non-enzymatic mechanism. In addition, both polysulfates and ZPGs induce proacrosin activation once they bind to the polysulfate-binding domain (PSBD) of the enzyme. We show here that the peptide (43)IFMYHNNRRYHTCGGILL(60) inhibited the proacrosin activation induced by either fucoidan or ZPGs. In addition, the peptide was recognized by the monoclonal antibody C5F10, which is directed against the PSBD region. Our data suggest that the PSBD is composed of many "subsites" that may or may not interact with each other.

Protease activity involvement in the passage of mammalian sperm through the zona pellucida

“…Binding of sulphate groups requires the correct presentation of positively charged basic residues of the surface of the proacrosin/acrosin. We have shown previously that model polysulfates such as fucoidan prevent activation of proacrosin into acrosin, a process that was mediated by a 18-residues peptide containin) a cluster of three basic amino acids (residues 59-76) (Moreno and Barros, 2000;Moreno et al, 2002). In this study we showed for the fi rst time that fucoidan and solubilized zona pellucida glycoproteins detach bound sperm from the zona pellucida, suggesting that the PSBD is indeed involved in the binding of acrosome-reacted spermatozoa to the zona.…”

“…The mechanism of interaction involves recognition of polysulphate groups on ZP glycoproteins by basic residues on the surface of proacrosin/acrosin, in a manner similar to that for heparin-anti-thrombin III (Moreno and Barros, 2000). Binding of sulphate groups requires the correct presentation of positively charged basic residues of the surface of the proacrosin/acrosin.…”

“…This polysulphated-binding domain (PSBD) must be different from the active site since neither specifi c inhibitors or mutations in the active-site amino acids affect acrosin interaction with those compounds (Lo Leggio et al, 1994;Moreno and Barros, 2000;Moreno et al, 1999;Moreno et al, 1998). Many biochemical and site-mutagenesis studies of different species have shown that the PSBD encompasses basic aminoacids (Crosby et al, 1998;Jansen et al, 1998;Moreno and Barros, 2000;Richardson and O'Rand, 1996).…”

“…This polysulphated-binding domain (PSBD) must be different from the active site since neither specifi c inhibitors or mutations in the active-site amino acids affect acrosin interaction with those compounds (Lo Leggio et al, 1994;Moreno and Barros, 2000;Moreno et al, 1999;Moreno et al, 1998). Many biochemical and site-mutagenesis studies of different species have shown that the PSBD encompasses basic aminoacids (Crosby et al, 1998;Jansen et al, 1998;Moreno and Barros, 2000;Richardson and O'Rand, 1996). In addition, an 18-amino acid peptide, containing those basic residues, is able to inhibit the activation of proacrosin induced by either fucoidan or solubilized zona pellucida glycoproteins (ZPG's) (Crosby et al, 1998;Jansen et al, 1998;Moreno and Barros, 2000;Richardson and O'Rand, 1996).…”

“…Many biochemical and site-mutagenesis studies of different species have shown that the PSBD encompasses basic aminoacids (Crosby et al, 1998;Jansen et al, 1998;Moreno and Barros, 2000;Richardson and O'Rand, 1996). In addition, an 18-amino acid peptide, containing those basic residues, is able to inhibit the activation of proacrosin induced by either fucoidan or solubilized zona pellucida glycoproteins (ZPG's) (Crosby et al, 1998;Jansen et al, 1998;Moreno and Barros, 2000;Richardson and O'Rand, 1996). The role of the second PSBD in sperm-zona pellucida binding during fertilization remains to be determined.…”