Interaction of Streptokinase and Human Plasminogen

Renzo, E.C. De; Boggiano, E.; Barg, William F.; Buck, Friedrich

doi:10.1016/s0021-9258(18)95979-x

Cited by 28 publications

(4 citation statements)

References 15 publications

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“…However, unlike other routinely used plasminogen activators (tissue plasminogen activator and urokinase), SK itself is not catalytically active. Rather, through a unique mechanism, SK binds and interacts with plasminogen to generate an active enzyme complex (Blatt et al, 1964;De Renzo et al, 1967;Reddy & Markus, 1972;Schick & Castellino, 1973). This enzyme, or "activator complex", then catalyzes the cleavage of the single-chain zymogen plasminogen to the active two-chain enzyme plasmin.…”

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confidence: 99%

Mutation of Lysines in a Plasminogen Binding Region of Streptokinase Identifies Residues Important for Generating a Functional Activator Complex

et al. 1996

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Through a unique but poorly understood mechanism, streptokinase (SK) interacts with human plasminogen to generate an "activator complex" that efficiently cleaves substrate plasminogen molecules. Previous studies have suggested that lysine residues in SK may play a role in the binding and function of the activator complex. To investigate this hypothesis, 10 different lysine residues in the plasminogen binding region of SK were altered to construct 8 recombinant (r) SK mutants. Only one double mutant, rSKK256,257A (replacing Lys with Ala at residues 256 and 257), showed a statistically significant reduction (63%) in binding affinity for Glu-plasminogen. This mutant also displayed a lagtime in the appearance of maximal activity, and modest impairments (2-5-fold) in kinetic parameters for amidolytic and plasminogen activator activity compared to rSK. In contrast, another mutant, rSKK332,334A, formed an activator complex with profound and nearly selective defects in the catalytic processing of substrate plasminogen molecules. When compared to rSK in kinetic assays of plasminogen activation, the rSKK332,334A mutant formed an activator complex that bound substrate plasminogens normally (normal K(m), but its ability to activate or cleave these molecules (kcat) was reduced by 34-fold. In contrast, in amidolytic assays, the kinetic parameters of rSKK332,334A showed only minor differences (< 2-fold) from rSK. Similarly, the binding affinity of this mutant to human Glu-plasminogen was indistinguishable from rSK [(2.6 +/- 0.8) x 10(9) vs (2.4 +/- 0.2) x 10(9) M-1, respectively]. In summary, these experiments have identified lysine residues in a plasminogen binding region of SK which appear to be necessary for normal high-affinity binding to plasminogen, and for the efficient catalytic processing of substrate plasminogen molecules by the activator complex.

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Mutation of Lysines in a Plasminogen Binding Region of Streptokinase Identifies Residues Important for Generating a Functional Activator Complex

et al. 1996

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“…The streptokinase activation of human plasminogen differs from typical enzymatic reactions in that streptokinase has a very low turnover number and in most investigations, including our own, the mole ratio of plasminogen to streptokinase does not exceed 200. The low turnover may be due to the slow dissociation of the plasmin-streptokinase complex which is readily isolated from mixtures of the two compounds (Davies et al, 1964;DeRenzo et al, 1967b).…”

Section: Discussionmentioning

confidence: 99%

“…Data to support this assumption have been obtained by indirect means such as using inhibitors and alternate substrates to demonstrate that the streptokinase/plasminogen ratio alters the esterolytic and caseinolytic characteristics of the mixture (Kline and Fishman, 1961;Markus, 1972, 1974; Markus and Werkheiser, 1964). The ability to isolate a stoichiometric complex of plasmin and streptokinase in the presence of the plasmin inhibitor e-aminocaproic acid (Davies et al, 1964;DeRenzo et al, 1967b) is also cited as evidence for the role of a stoichiometric complex in the activation of human plasminogen. The decreased caseinolytic activity of plasmin in the presence of large amounts of streptokinase may be explained by alternate substrate inhibition since plasmin is known to cleave various peptide bonds of strepf Contribution No.…”

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Kinetic mechanism of the activation of human plasminogen by streptokinase

Kosow¹

1975

Biochemistry

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A method of determining the initial rate of plasminogen activation has been developed. The method has been used to investigate the mechanism of activation of human plasminogen by streptokinase. Plasmin formation follows saturation kinetics. Inhibition of plasmin formation by epsilon-aminocaproic acid is uncompetitive with a Ki of 0.6 mM. A model consistent with the data is that streptokinase induces a conformational change in the plasminogen molecule, producing an active center which cleaves an internal peptide bond to produce plasmin. Thus, streptokinase functions as a catalytic allosteric effector.

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“…Kinetic parameters have been determined for both the Plg-SK and Pln-SK enzyme complexes (Buck & Boggiano, 1971; Reddy & Markus, 1974;Wohl et al, 1977Wohl et al, -1980Morris et al, 1981). Various methods have been used to dissociate the Plg-SK enzyme complex in an effort to study the individual moieties (De Renzo et al, 1967;Buck et al, 1968;Summaria et al, 1968Summaria et al, , 1971Brockway & Castellino, 1974), but a pure, structurally intact Pig moiety, with or without an active site, had not been pre-preparation were determined on two synthetic chromogenic substrates. With H-D-valyl-L-leucyl-L-lysyl-p-nitroanilide, a Km of 417 /tM and a A:cat of 8.44 s_1 were obtained; with Tos-glycyl-L-prolyl-L-lysyl-p-nitroanilide, a Km of 307 juM and a t of 12.83 s_1 were obtained.…”

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confidence: 99%