KINETICS OF ACTIVATION OF VARIANT PLASMINOGENS IN A NONCROSSLINKED FIBRIN CLOT - ABNORMAL PLASMIN GENERATION WITH THE PLASMINOGEN - STREPTOKINASE COMPLEX, UROKINASE AND TISSUE-PLASMINOGEN ACTIVATOR

A purified clot lysis system (noncrosslinked) was developed to evaluat e and study variant plasminogens from patients with a history of throm bophilia. Plasmin generation was studied in the soluble phase and in t he clot lysis system with different activators, streptokinase and its derivatives, urokinase and tissue plasminogen activator. Five of the v ariant plasminogens (D1, D2, K, S, and F) generated low levels of plas min in the soluble phase, whereas two (M and Y) generated near normal levels of plasmin. Plasmin generation in the clot lysis system divided the variants into three groups: one variant (D1) had a very prolonged lysis time or a negligible plasmin generation rate with three direct activators, three variants (D2, K and S) had moderately prolonged lysi s times or low plasmin generation rates with these activators, and thr ee variants (F, M, and Y) had normal lysis times and normal plasmin ge neration rates with these activators. Kinetics of activation parameter s with three direct plasminogen activators were determined from a non- linear regression analysis of plots of velocity versus plasminogen con centration and were verified by Lineweaver-Burk plots where possible. With plasminogen.streptokinase, the four variants with prolonged lysis times, had normal apparent Michaelis constants and low catalytic rate constants. Three variants with normal lysis times had high apparent M ichaelis constants showing decreased affinity for the plasminogen.stre ptokinase activator complex, and high catalytic rate constants. With u rokinase, the variant with the longest clot lysis time had a high appa rent Michaelis constant and a low catalytic rate constant, three varia nts with prolonged lysis times gave low catalytic rate constants; two of them gave normal apparent Michaelis constants and one gave a high a pparent Michaelis constant. Of the three variants with normal lysis ti mes, one gave normal kinetic constants and two had high apparent Micha elis constants with high catalytic rate constants. With tissue plasmin ogen activator, the variant with the longest clot lysis time gave a hi gh apparent Michaelis constant and a high catalytic rate constant. The three variants with prolonged lysis times gave variable results, one had a slightly higher apparent Michaelis constant with a low catalytic rate constant, the second one had a very low apparent Michaelis const ant with a low catalytic rate constant and the third one had a high ap parent Michaelis constant with a normal catalytic rate constant. Three variants with normal lysis times had slightly higher Michaelis consta nts and near normal catalytic rate constants. With the noncrosslinked fibrin substrate, all of the variant plasminogens have lowered catalyt ic efficiencies with two or three plasminogen activators. The theoreti cal aspects of these plasmin generation data are discussed.