Interaction Mechanism of Thrombin with Functional Polystyrene Surfaces: A Study Using High-Performance Affinity Chromatography

Abstract

Insoluble sulfonated polystyrenes (PSSO 3 ) and their L-arginyl methyl ester derivatives ( PAOM) have beeninvestigated by high-performance affinity chromatogra-phy.Highly active thrombin(Th) was found to be strongly adsorbed on both surfaces and was eluted by increasing ionic strength of the eluent. Compared to the PSSO 3 surface, desorption from the PAOM resin occurred at a lower salt concentration and with a more specific enzyme-resin interaction. Masking of the enzyme's active site by its natural inhibitor antithrombin III (AT III) caused a total disappearence of its affinity for both types of surface. It was demonstrated that the active seryl residue of thrombin was involved in enzyme-PAOM interactions. In contrast, this same site of the enzyme was always available after binding to PSSO 3 resin. This allowed a biospecific desorption procedure using AT III, heparin (Hep) and Hep-AT III complex. The present work provides further confirmation of the "AT III-like" and "heparin -like" properties, respectively, of PAOM and PSSO 3 materials.