A method for characterising serum fibrinogen and fibrin degradation products
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Agarose
Agarose gel electrophoresis
Polyacrylamide
Degradation
Human plasminogen was treated with either DFP, TLCK, or SBTI under conditions which completely inhibited the plasmin contaminant in our preparation. Each treated plasminogen preparation could be completely activated with low molar ratios of streptokinase to plasminogen in 24 hr, in 25% glycerol. Activation of DFP-treated plasminogen with either streptokinase or a DFP-treated equimolar human plasmin-streptokinase complex could also be carried out in 10-2M DFP, in 25% glycerol; however, much longer periods of time were required to obtain nearly complete activation. Under these conditions for activation, the plasmin contaminant in our human plasminogen preparation is therefore not essential for streptokinase-activation of the proenzyme. These data indicate that streptokinase, alone or in the complex, acts directly on human plasminogen. A new electrophoretic method for differentiating human plasminogen from human plasmin is described.
Fibrinolysin
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Summary The novel mutant streptokinase, SK-K59E, can activate human plasminogen as efficiently as the purified commercially available streptokinase. Several peptide bonds including Lys59-Ser60 in native streptokinase were hydrolyzed in reaction with plasmin and peptides of small molecular masses were generated. The plasminogen activator activity of native streptokinase in reaction with human plasmin declined to 25% of the original activity in a 120-min incubation. On the other hand, the NH2-terminal peptide of SK-K59E remained intact in reaction with plasmin and the activator activity of streptokinase decreased to 75% of the original activity in 120 min. The major degraded peptide fragments of native streptokinase in reaction with plasmin had molecular masses of 36 and 30 kDa. However, two major peptide fragments of 42 and 34 kDa were observed in the reaction of SK-K59E with human plasmin. The 42 kDa peptide fragment, which contained NH2-terminal of streptokinase, could activate human plasminogen as efficiently as the native streptokinase. SK-K59E can induce greater degree of caseinolysis and fibrinolysis than the native streptokinase. In conclusion, the results demonstrate that the prevention of cleavage at Lys59 of streptokinase prolongs the half-life of streptokinase in complex with plasmin and that the NH2-terminal of streptokinase (Ile1-Lys59) plays an important role in maintaining its stability.
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Journal Article A Role For Fibrinogen In The Streptokinase-Dependent Acquisition Of Plasmin(ogen) By Group A Streptococci Get access Hong Wang, Hong Wang Search for other works by this author on: Oxford Academic PubMed Google Scholar Richard Lottenberg, Richard Lottenberg Search for other works by this author on: Oxford Academic PubMed Google Scholar Michael D. P. Boyle Michael D. P. Boyle Reprints or correspondence: Dr. Michael D. P. Boyle, Dept. of Microbiology, Medical College of Ohio, P.O. Box 10008, Toledo, OH 43699-0008. Search for other works by this author on: Oxford Academic PubMed Google Scholar The Journal of Infectious Diseases, Volume 171, Issue 1, January 1995, Pages 85–92, https://doi.org/10.1093/infdis/171.1.85 Published: 01 January 1995 Article history Received: 20 April 1994 Revision received: 12 August 1994 Published: 01 January 1995
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Summary Procedures have been described for the purification of streptokinase and human plasmin, and molecular weights of 42,600 and 88,800 respectively have been determined. Equilibration of streptokinase with plasmin solutions produced an inhibition of caseinolytic activity and indicated a mole-mole interaction. This inhibitory activity was lost following acid treatment. When these data are considered with the previously reported findings using plasminstreptokinase mixtures as plasminogen activators, it would appear that plasmin and streptokinase react to form a complex which has the ability to convert both human and bovine plasminogen to plasmin.
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Plasminogen activation to plasmin is due to enzymatic cleavage of a single peptide bond in the zymogen molecule. Streptokinase is not an enzyme and its activation of plasminogen is indirect. Streptokinase forms stoichiometric complexes with plasmin and plasminogen and these complexes activate plasminogen to plasmin. Streptokinase is species selective in its action.
Zymogen
Cleavage (geology)
Fibrinolysin
Peptide bond
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Fibrinolysin
Fibrinolytic agent
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Aprotinin
Dissociation constant
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Abstract DNA electrophoresis in gels and solutions of agarose and polyacrylamide was objectively evaluated with regard to separation efficiency at optimal polymer concentrations. In application to DNA fragments, polyacrylamide gels were superior for separating fragments of less than 7800 bp, and agarose gels are the best choice for larger fragments. Agarose solutions are nearly as good as polyacrylamide gels for small DNA (< 300 bp). Agarose solutions have a higher efficiency than polyacrylamide solutions for DNA of less than 1200 bp. Separation efficiency sharply decreases with increasing length of DNA. Retardation in polyacrylamide solutions was found to depend on polymer length in a biphasic fashion. The choice of resolving polymer concentrations depends on the progressive stretching of DNA in proportion to polymer concentration. The rate of that stretching appears higher in polyacrylamide solution than in gels or in liquid or gelled agarose. Application of polymer solutions to capillary electrophoresis raises further problems concerning agarose plugs, DNA interactions with the polymers, operation at low field strength and long durations as well as detection sensitivity.
Polyacrylamide
Agarose
Agarose gel electrophoresis
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