Summary
Human antithrombin III was purified from fresh human plasma by affinity chromatography on heparin-Sepharose®, affinity chromatography on concanavalin A Sepharose®, gel filtration on Ultrogel® AcA 34, ion exchange chromatography on DEAE A-50 Sephadex® and preparative agarose gel electrophoresis. The hydrolytic activity of urokinase (plasminogen activator from urine) on acetyl-glycyl-L-lysine methyl ester acetate (Ac-gly-lys-OMe Ac) was inhibited by antithrombin III in a slow time-dependent manner. Heparin accelerated the reaction between activator and inhibitor. Inhibition of catalytic activity was associated with the formation of an 1:1 molar complex between activator and inhibitor as revealed by sodium dodecyl sulphate polyacrylamide gel electrophoresis. The complex was also demonstrated by crossed Immunoelectrophoresis against anti-antithrombin III.
