Enhancement of the streptokinase-catalyzed activation of human plasminogen by human fibrinogen and its plasminolysis products

Abstract

The effects of human fibrinogen, and several plasmin-derived fragments of fibrinogen, on the streptokinase-induced activation of human plasminogen (Pg) have been investigated. Fibrinogen stimulates the rate of activation of human Glu1-Pg, Lys77-Pg, and Val442-Pg. The cofactor activity of fibrinogen appears to reside mainly in the D-domain region, since purified fragment D is active in this system. Fibrinogen fragment E was not active in this regard. The cofactor activity of fragment D was partially dependent on the presence of Ca2+. This effect of Ca2+ was likely due to its stabilizing influence on fragment D, as revealed by studied employing differential scanning calorimetry. Conversion of fragment D1 to fragments D2--5 did not alter the cofactor activity. Steady-state kinetic analysis of the activation of Val442-Pg by the streptokinase-Val442-plasmin complex demonstrated that the Km decreased approximately 2-fold, in the presence of fragment D1. Very little change in the steady-state kinetic parameters for Glu1-Pg and Lys77-Pg, when activated by the streptokinase-Lys77-plasmin complex, was noted in the presence of fragment D1. It was also found that both fibrinogen and fibrinogen fragment D1 increased the rate of formation of the active site in the streptokinase-plasminogen complex, of all forms of plasminogen, and that this effect was sufficient to explain the overall stimulation of the activation of plasminogen by fibrinogen and fibrinogen fragment D1.

Kinetic Analyses of Potentiation of Plasminogen Activation by Streptokinase in the Presence of Fibrin or Its Degradation Products

“…The observation that the activity of the SKplg complex was enhanced in the presence of fibrin degradation products generated by vascular plasminogen activator released in the plasma by venous stasis [15] pointed to the clinical importance of these potentiating agents. Kinetic analyses and active site titra tion indicated that potentiating agents accel erated the appearance of an active site in the SK-plg complex, thus resulting in the faster formation of SK-plg' complex [4]. Previous ly, we have noticed that the hydrolysis of S-2251 by the mixture of plasminogen and SK took place linearly for some time intervals after an initial lag period [11], These results indicate that the SK-plg complex primarily hydrolyzes S-2251 at low concentrations of SK, and that the conversion of the remaining plasminogen to plasmin by the SK-plg com plex was less effective than the hydrolysis of S-2251.…”

Analogs of Human Plasminogen That Are Labeled with Fluorescence Probes at the Catalytic Site of the Zymogen

“…Non-proteolytic activation of Pg by SK is closely coupled with enzymatic activation of Pg by Pg-SK and plasmin-SK complexes, by their cleavage of the Arg 561 -Val 562 bond (Bajaj and Castellino, 1977;GonzalezGronow et al, 1978;Davidson et al, 1990). Similar to activation by the physiological proteinases, the rate of SK-initiated plasmin formation is regulated by interactions with fibrin (Strickland et al, 1982;Chibber et al, 1985;Cassels et al, 1987), Pg conformational equilibria (Chibber and Castellino, 1986), and plasmin proteolytic reactions (Strickland et al, 1982). Studies aimed at defining the molecular mechanisms of Pg activation have been complicated by the concurrent plasmin-catalyzed reactions.…”

Control of Human Plasminogen Activation

“…It has been established that the SK-induced activation rate of human Glu,Pg is stimulated by fibrinogen (8)(9)(10)20) and inhibited by Cl" (14)(15)(16). We have shown that both effectors function by the respective stimulation (20) or inhibition (15) Here, SK and HPg interact to form a stoichiometric complex (SK-HPg), from which, most likely via conformational altera tions, a complex, SK-HPg*, is formed, that possesses plasminogen activator activity.…”

“…Fibrinogen, fibrin and degradation products of these proteins function as positive effec tors of plasminogen activation with tPA and SK as activators (7)(8)(9)(10). On the other hand, anions inhibit the activation of human plasminogen with all of the above activators (11)(12)(13)(14)(15)(16).…”