A BATTERY OF MONOCLONAL-ANTIBODIES THAT INDUCE UNIQUE CONFORMATIONS TO EVOLVE CRYPTIC BUT CONSTITUTIVE FUNCTIONS OF PLASMINOGEN

Two groups of anti-plasminogen monoclonal antibodies, whose epitope wa s either in the kringle 1+2+3 domain (F3P2, F11P5, F11P6, and F12P18) or the kringle 5 domain (F1P6 and F12P16), were isolated and their eff ects on the conformation of plasminogen were explored. All antibodies except F1P6 had 3- to 10-fold higher affinity toward Lys-plasminogen t han Glu-plasminogen. F1P6 exhibited a comparable affinity to Glu- and Lys-plasminogen. Among these, only F11P5 binding was inhibited by epsi lon-amino-n-caproic acid (EACA) in a concentration-dependent manner, w ith half maximal inhibition at 3 mM. From a competition assay, we conc luded that the epitopes of F11P5, F11P6, and F12P18 should be very clo se, and located at or near the low affinity lysine binding site on the kringle 2+3, These three antibodies dramatically enhanced the binding of Glu-plasminogen to the other antibodies, except to F1P6. Interesti ngly, F3P2, whose non-overlapping epitope was in the kringle 2+3 domai n, also augmented the binding of Glu-plasminogen to the other antibodi es. In contrast, we did not observe enhanced binding of Lys-plasminoge n to one antibody in the presence of the other antibodies, and the bin ding of Glu-plasminogen to these antibodies did not increase in the pr esence of 10 mM EACA, In the presence of these antibodies, including F 1P6, Glu-plasminogen bound more efficiently to immobilized degraded fi brin, with a binding profile similar to Lys-plasminogen, All antibodie s except F1P6 enhanced the conversion rate of plasminogen to plasmin r emarkably. Taken together, we propose that these two groups of monoclo nal antibodies can dissociate the intramolecular interactions of Glu-p lasminogen and induce the conformational transition of Glu-plasminogen to Lys-plasminogen. In addition, the kringle 2+3 and kringle 5 struct ures of Glu-plasminogen liganded with EACA are distinct from the Lys-p lasminogen structure.