Binding of recombinant human proacrosin/acrosin to zona pellucida glycoproteins. II. Participation of mannose residues in the interaction.

To assess the interaction of human proacrosin/acrosin with mannose residues coupled to a protein backbone. Prospective study. Basic research laboratory. Recombinant proteins derived from human proacrosin (Rec-40, Rec-30, Rec-20, Rec-10, and Rec-6) and bovine serum albumin (BSA)-mannose as ligand. In vitro binding assay developed to assess proacrosin/acrosin-BSA-mannose interaction. Proacrosin/acrosin binding to BSA-mannose; estimation of binding affinity. All recombinant proteins of acrosin but Rec-6 (residues 1-59 of proacrosin) specifically bound to BSA-mannose. Rec-40 (proacrosin) showed the highest binding affinity (dissociation constant K(d), 162 nM), followed by N-terminal fragments Rec-30 (248 nM), Rec-20 (359 nM), and Rec-10 (402 nM). A significant decrease in binding activity was observed when acrosin proteins were treated with denaturing agents such as urea or heat. The beta-mercaptoethanol treatment produced a 39% decrease on Rec-30 binding to BSA-mannose; in contrast, no effect was observed with Rec-40, suggesting the presence of at least two types of mannose-binding sites. [1] Proacrosin interacts with mannose residues through binding sites located at both the N- and C-terminal portion of the protein, [2] the full-length protein is required for maximal BSA-mannose binding, and [3] binding sites are stabilized by noncovalent bonds and by disulfide linkages.