Presence in bovine milk of two protease inhibitors of the plasmin system.

Proteolysis, caused by the serine proteinase plasmin (EC 3.4.21.7) that is present in milk, influences the quality of dairy products. Within the plasmin system, activators and inhibitors control plasmin activity. This study investigated the presence in bovine milk of two serine proteinase inhibitors of the plasmin system, alpha(2)-antiplasmin and plasminogen activator inhibitor-1, and an isolation procedure used for partial purification of them from milk. Two colorimetric assays were used to detect either plasmin inhibitor activity or plasminogen activator inhibitor activity. Two inhibitors were partially purified from milk using a combination of ammonium sulfate fractionation and concanavalin A affinity chromatography. Plasminogen activator inhibitor-1 and alpha(2)-antiplasmin antigens, which were associated with the inhibitory activities from bovine milk, were visualized by Western blot using commercial polyclonal antibodies raised against the corresponding human inhibitors. Both inhibitors were present in milk as several forms, possibly from the formation of complexes with other milk proteins. The predominant forms of the inhibitors in milk exhibited an approximate molecular mass of 60 kDa for alpha(2)-antiplasmin and 55 kDa for plasminogen activator inhibitor-1.