Direct Activation of Human Plasminogen by Streptokinase

Summaria, Louis; Hsieh, Betty; Groskopf, William R.; Robbins, Kenneth C.

doi:10.3181/00379727-130-33645

Cited by 26 publications

(8 citation statements)

References 3 publications

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“…Induction of such a conformational change in the absence of activation cleavage and concomitant generation of the mature amino terminus is particularly intriguing but not unprecedented. Similar activation of plasminogen by binding to streptokinase (42)(43)(44)(45)(46)(47) as well as activation of prothrombin by binding to staphylocoagulase (48,49) has been described previously. Although the mechanism of this nonclassical, nonproteolytic activation of serine protease zymogens remains completely unclear, the behavior of single-chain t-PA/R15E,D194E and t-PA/R15E,D194N suggests that Asp-194 does not play an essential role in the process.…”

Section: Discussionsupporting

confidence: 69%

Variants of Tissue-type Plasminogen Activator with Substantially Enhanced Response and Selectivity toward Fibrin Co-factors

Strandberg

Madison

1995

Journal of Biological Chemistry

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Unlike most proteases, tissue-type plasminogen activator (t-PA) is not synthesized as an inactive precursor or zymogen. Instead, the single-chain "proenzyme" form of t-PA possesses very significant catalytic activity. Recent investigations of the molecular basis of the unusually high enzymatic activity of single-chain t-PA have focused attention upon Asp-194, a residue that is invariant among chymotrypsin-like enzymes. The critical role of this residue in securing the active conformation of mature chymotrypsin-like enzymes has been discussed extensively. Subsequent work, however, has indicated that this conserved residue can also form interactions that dramatically influence the catalytic activity of serine protease zymogens. While Asp-194 forms interactions that suppress the activity of the zymogen chymotrypsinogen, it may, by contrast, directly promote the catalytically active conformation of single-chain t-PA. To test the hypothesis that Asp-194 promotes the activity of both single-and two-chain t-PA and therefore plays opposing roles in single-chain t-PA and chymotrypsinogen, and also to examine whether this invariant residue plays an essential role in the stimulation of t-PA by fibrin, we used site-directed mutagenesis to construct the following variants of t-PA: t-PA/D194E, t-PA/ D194N, t-PA/R15E,D194E, and t-PA/R15E,D194N. In the absence of fibrin, the activity of enzymes carrying a mutation at position 194 was reduced by factors of 1000 -2000 compared to wild type t-PA. Similar reductions of activity were observed for both single-and two-chain variants, suggesting an important role for Asp-194 in both forms of the enzyme. The mutated enzymes, however, displayed a dramatically enhanced response to fibrin monomers. While the activity of wild type t-PA was stimulated by fibrin monomers by a factor of 960, the corresponding stimulation factor for the mutated enzymes varied from 498,000 -1,050,000.Many critical biological processes (1-3) depend on specific cleavage of individual target proteins by serine proteases. One important example is the dissolution of blood clots in which the initiating and rate-limiting step is the activation of the circulating zymogen plasminogen by tissue-type plasminogen activator (t-PA) 1 (4, 5).Unlike typical chymotrypsin-like enzymes, the single-chain or "proenzyme" form of t-PA possesses high catalytic activity (6 -12). In the absence of the co-factor fibrin, single-chain t-PA is approximately 8% as active as two-chain t-PA. In the presence of fibrin, however, single-and two-chain t-PA display equivalent enzymatic activity. "Zymogen activation" of singlechain t-PA, therefore, can be accomplished either by activation cleavage or by binding to the co-factor fibrin.The unusually high, intrinsic enzymatic activity of singlechain t-PA may reflect both the absence of interactions, present in typical proenzymes, that suppress activity of the zymogen and the presence of interactions, absent in typical zymogens, that stabilize an active conformation of the single-chain enzyme. Recent st...

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Section: Discussionsupporting

confidence: 69%

Variants of Tissue-type Plasminogen Activator with Substantially Enhanced Response and Selectivity toward Fibrin Co-factors

Strandberg

Madison

1995

Journal of Biological Chemistry

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“…A complex is also produced by the interaction of streptokinase and human plasminogen, but it is felt that the plasminogen moiety in the complex may have to be altered in some way or to be transformed into plasmin in order for the complex to acquire activity (6,21,25). Finally, streptokinase has been shown to be able to activate human plasminogen directly (22), and it is possibly in this manner that the first plasmin is formed in the human plasma clot lysis test. Once a trace of plasmin is present, the stage is set for activator formation and thus for much more rapid further activation.…”

Section: Discussionmentioning

confidence: 99%

Inhibition of Coagulation and Fibrinolysis by Aromatic Amidino Compounds

Geratz¹

1971

Thromb Haemost

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Summary1. Aromatic diamidines which are potent inhibitors of trypsin possess a marked inhibitory effect on the clotting activity of human thrombin and on the prothrombin time and partial thromboplastin time of human plasma. They also block the contact activation phase of the coagulation process. The strongest inhibitor among the compounds tested was M & B 4596 which was followed in second place by pentamidine.2. Pentamidine was 10 times more active than ε-ACA in impeding streptokinase-induced lysis of human plasma clots. It was 100-200 times stronger than ε-ACA in inhibiting the activation of bovine plasminogen by activators formed from the interaction between streptokinase and either human plasmin(ogen) or human plasma.3. The prothrombin time and partial thromboplastin time of canine plasma were less susceptible to inhibition by pentamidine than the same tests on human plasma. Clot lysis in the canine system was inhibited by pentamidine to a similar degree as in the human system. After intravenous injection of pentamidine in the dog there occurred the expected prolongation of the partial thromboplastin time and of the clot lysis time.

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“…This interaction leads to formation of a modified streptokinase capable of directly activating human plasminogen. However, several authors have shown conclusively that preformed human plasmin was not a requirement for the generation of the activator active site (Mounter and Shipley, 1958;Summaria et al, 1969;McClintock and Bell, 1971;Reddy and Markus, 1972). This fact was not discussed in the mechanism proposed by Taylor and Beisswenger (1973).…”

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confidence: 99%

Interaction of streptokinase and rabbit plasminogen

Schick¹

1973

Biochemistry

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The ability of rabbit plasminogen and plasmin to form complexes with streptokinase has been studied and compared to the human system. At 1 : l or 1O:l ratios of streptokinase to rabbit plasminogen, no complex was observed by sucrose density centrifugation; however, almost all the rabbit plasminogen was converted to plasmin. Under identical conditions with human plasminogen, a complex was formed which consisted of altered streptokinase and human plasmin. When the acylating agent p-nitrophenyl p'-guanidinobenzoate was added to either human or rabbit plasminogen prior to addition of a molar equivalent of streptokinase, reactive complexes were formed in each case which consisted of human or rabbit plasminogen and native streptokinase. Upon treatment of human plasmin with a molar equivalent of streptokinase, a complex is formed which consists of altered streptokinase and human plasmin. This complex possesses the ability to activate P lasminogen is a single-chain protein of mol wt 81,000-88,000, depending on the species (Barlow et al., 1969; Sodetz et ul., 1972), and plasmin is a two-chain molecule of approximately the same molecular weight, stabilized by disulfide bond(s) (Barlow et ul., 1969;Sodetz et al., 1972). Many agents mediate the conversion of plasminogen to plasmin, among which is the bacterial endotoxin streptokinase. The mechanism of activation of human plasminogen by streptokinase has been a widely studied subject. The problem resolves itself into the manner in which the nonproteolytic protein, streptokinase, catalyzes the proteolytic cleavage required for conversion of plasminogen into plasmin. Further, the fact that not all species of plasminogen are capable of activation by streptokinase raises other interesting questions.

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Direct Activation of Human Plasminogen by Streptokinase

Cited by 26 publications

References 3 publications

Variants of Tissue-type Plasminogen Activator with Substantially Enhanced Response and Selectivity toward Fibrin Co-factors

Variants of Tissue-type Plasminogen Activator with Substantially Enhanced Response and Selectivity toward Fibrin Co-factors

Inhibition of Coagulation and Fibrinolysis by Aromatic Amidino Compounds

Interaction of streptokinase and rabbit plasminogen

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