THE SOLUTION-PHASE INTERACTION BETWEEN APOLIPOPROTEIN(A) AND PLASMINOGEN INHIBITS THE BINDING OF PLASMINOGEN TO A PLASMIN-MODIFIED FIBRINOGEN SURFACE

the present study, we assessed the binding of recombinant forms of apo lipoprotein(a) [r-apo-(a)] to plasminogen. Apo(a)-plasminogen interact ions were demonstrated to be lysine-dependent, as they were abolished by the addition of E-aminocaproic acid. Binding of r-apo(a) and plasma -derived Lp(a) to Glu-plasminogen was assessed in solution using a mut ant form of recombinant plasminogen [Plg(S741C)] labeled at the active site with 5'-(iadoacetamido)fluorescein. High-affinity binding of apo (a) to plasminogen was observed with the 17-kringle r-apo(a) (K-d = 20 .1 +/- 3.3 nM) as well as with plasma-derived Lp(a) (K-d = 5.58 +/- 0. 08 nM). Binding studies using various truncated and mutant forms of r- apo(a) demonstrated that sequences within apo(a) kringle IV types 2-9 and the strong lysine binding site (LBS) in apo(a) kringle IV type 10 are not required for high-affinity binding to plasminogen. In all case s, the binding stoichiometry for the apo(a)-plasminogen interaction wa s determined to be 1:1. Binding data obtained using a 17-kringle r-apo (a) derivative lacking the protease-like domain (17K Delta P; K-d = 31 58 +/- 138 nM) indicate that sequences within the protease-like domain of apo(a) mediate its interaction with LBS in plasminogen. We determi ned that r-apo(a) and plasminogen bind to distinct sites on plasmin-mo dified fibrinogen with the concentration of plasminogen binding sites exceeding the concentration of r-apo(a) sites by a factor of 10, Furth ermore, r-apo(a) is capable of inhibiting the binding of plasminogen t o plasmin-modified fibrinogen surfaces, an effect which we show is att ributable to the formation of a solution phase apo(a)/plasminogen comp lex which exhibits a greatly reduced affinity for plasminogen binding sites on plasmin-modified fibrinogen, The results of this study provid e new insights into the mechanism by which apo(a) and Lp(a) may inhibi t fibrinolysis, thus contributing to the atherothrombotic risk associa ted with this lipoprotein.