Interaction of streptokinase with plasminogen. Isolation and characterization of a streptokinase degradation product.

G E Siefring,F J Castellino

doi:10.1016/s0021-9258(17)33335-5

Abstract

When streptokinase is incubated with human or rabbit plasminogen, one event which occurs is a specific fragmentation of streptokinase. At least five major identifiable streptokinase fragments appear with time, and they possess molecular weights of approximately 40,000 (SK 1), 36,000 (SK 2), 31,000 (SK 3), 26,000 (SK 4), and 10,000 (SK 5) under denaturing conditions, as observed on calibrated sodium dodecyl sulfate-polyacrylamide gels, compared to native streptokinase of molecular weight 45,000. The amount of each of the fragments generated at given times of incubation of plasminogen and streptokinase depends upon the species of plasminogen employed. Utilizing rabbit plasminogen and streptokinase, the SK 4 fragment was purified. This fragment arises by proteolysis at both the NH2 and COOH regions of native streptokinase. However, when isolated utilizing dilute aqueous buffers, the SK 4 fragment contained a portion of the original NH2 terminus of native streptokinase noncovalently bound to the molecule (SK 4'). SK 4' is capable of activating human plasminogen to plasmin, albeit more slowly than native streptokinase. However, the SK 4'-human plasmin complex possess only very weak plasminogen-activating activity toward sheep plasminogen. Upon removal of the noncovalently bound small NH2-terminal peptide of native streptokinase from SK 4', SK 4 is formed. This particular fragment possesses practically no human plasminogen-activating activity and cannot be used as an activator of sheep plasminogen, even with added human plasminogen.

Highlights

When streptokinase is incubated with human or rabbit plasminogen, one event which occurs is a specific fragmentation of streptokinase
At least five streptokinase fragments can be readily observed with rabbit plasminogen
Since plasmin alone will not convert plasminogen to plasmin, it is reasonably assumed that the integrity of streptokinase in the complex is necessary for plasminogen activation

Summary

Introduction

When streptokinase is incubated with human or rabbit plasminogen, one event which occurs is a specific fragmentation of streptokinase. Utilizing rabbit plasminogen and streptokinase, the SK 4 fragment was purified. When isolated utilizing dilute aqueous buffers, the SK 4 fragment contained a portion of the original NH, terminus of native streptokinase noncovalently bound to the molecule (SK 4’). Upon removal of the noncovalently bound small NH,-terminal peptide of native streptokinase from SK 4’, SK 4 is formed. This particular fragment possesses practically no human plasminogen-activating activity and cannot be used as an activator of sheep plasminogen, even with added human plasminogen. The mechanism of activation of human plasminogen by streptokinase is complex, and occurs indirectly via a plasminogen activator complex (l-3). Portions of this work were taken from a thesis submitted to the Graduate School of the University of Notre Dame, in partial fulfillment of the requirements for the Ph.D. degree

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