Aggregated Forms of Bull Seminal Plasma Proteins and Their Heparin-Binding Activity

Abstract

Heparin-binding activity of bull seminal plasma proteins was shown to be dependent on their aggregation state. The protein fraction interacting with immobilized heparin was characterized by large polydispersity in the region of molecular weight of 60 000-10 000, while that not retained on the affinity carrier was present as aggregates with molecular weight >100 000. Components of heparin-binding and non-heparin-binding fractions were separated by RP HPLC (reversed-phase HPLC) and analyzed by SDS (sodium dodecyl sulfate) electrophoresis and N-terminal sequencing. Size exclusion chromatography of whole seminal plasma and heparin-binding proteins in the presence of D-fructose (as a component of seminal plasma) showed that the region of molecular weights of protein-associated forms was shifted to lower values. An increase of heparin-binding activity of bull proteins, as determined by ELBA (Enzyme-Linked Binding Assay), correlates with a decrease of their aggregation state. The modulation of the aggregation state of bull proteins by seminal plasma components and, in this way, also of their heparin-binding properties suggests possible mechanisms for capacitation mediated by these proteins.