Secondary-site binding of Glu-plasmin, Lys-plasmin and miniplasmin to fibrin

Abstract

Active-site-inhibited plasmin was prepared by inhibition with D-valyl-L-phenylalanyl-L-lysylchloromethane or by bovine pancreatic trypsin inhibitor (Kunitz inhibitor). Active-site-inhibited Glu-plasmin binds far more strongly to fibrin than Glu-plasminogen [native human plasminogen with N-terminal glutamic acid (residues 1-790)]. This binding is decreased by .alpha.2-plasmin inhibitor and tranexamic acid and is, in the latter case, related to saturation of a strong lysine-binding site. In contrast, .alpha.2-plasmin inhibitor and tranexamic acid have only weak effects on the binding of Glu-plasminogen to fibrin. This demonstrates that its strong lysine-binding site is of minor importance to its binding to fibrin. Active-site-inhibited Lys-plasmin and Lys-plasminogen (Glu-plasminogen lacking the N-terminal residues Glu1-Lys76, Glu1-Arg67 or Glu1-Lys77) display binding to fibrin similar to that of active-site inhibited Glu-plasmin. Tranexamic acid or .alpha.2-plasmin inhibitor similarly decrease their binding to fibrin. Glu-plasminogen and active-site-inhibited Glu-plasmin have the same gross conformation and conversion into their respective Lys-forms produces a similar marked change in conformation. This change is not essential to the degree of binding to fibrin or the effect of .alpha.2-plasmin inhibitor and tranexamic acid on this binding. The conversion of miniplasminogen (Glu-plasminogen lacking the N-terminal residues Glu1-Val441) into active-site-inhibited miniplasmin makes no difference to the degree of binding to fibrin, which is similarly decreased by the addition of tranexamic acid and unaffected by .alpha.2-plasmin inhibitor. Active-site-inhibited Glu-plasmin, Lys-plasmin and miniplasmin have lower fibrin-binding values in a plasma system than in a purified system. Results with miniplasmin(ogen) indicate that plasma proteins other than .alpha.2-plasmin inhibitor and histidine-rich glycoprotein decrease the binding of plasmin(ogen) to fibrin.