Fibrin Suspension as a Substrate for Plasmin: Determination and Kinetics

Summary

Fibrin polymers formed from fibrinogen with thrombin in the presence of EDTA were suspended in a medium containing glucose, arabic gum and imidazole-HCl buffer and were sonicated at 20 kHz for 20 min to make a suspension containing fibrin particles of small size. The fibrin suspension was used as a substrate of plasmin for determining the enzymic activity of plasmin and plasminogen activated with urokinase. The kinetic study on the reaction of the fibrin particles with plasmin in the presence and the absence of fibrinogen revealed that Km value of fibrin for plasmin is 4.2 × 10^-7 M and the Ki value of fibrinogen is 1.2 × 10^-5 M.

• References

• 1 Alkjaersig N, Fletcher AP, Sherry S. 1959; The mechanism of clot dissolution by plasmin. J. Clin. Invest 38: 1086-1095
• 2 Aster RH, Jandl JH. 1964; Platelet sequestration in man. I. Methods. J. Clin. Invest 43: 843-855
• 3 Astrup T, Mullertz S. 1952; The fibrin plate method for estimating fibrinolytic activity. Arch. Biochem. Biophys 40: 346-351
• 4 Genton E, Fletcher AP, Alkjaersig N, Sherry S. 1964; Assay of plasma thrombolytic activity with fluorescein-labelled clots. J. Lab. Clin. Med 64: 313-320
• 5 Lassen M. 1952; Heat denaturation of plasminogen in the fibrin plate method. Acta Physiol. Scand 27: 371-376
• 6 Lowry OH, Rosebrough NJ, Farr AL, Randall RJ. 1951; Protein measurement with the folin phenol reagent. J. Biol. Chem. 193: 265-275
• 7 Marder VJ, Shulman NR. 1969; High molecular weight derivatives of human fibrinogen produced by plasmin. J. Biol. Chem 244: 2111-2119
• 8 Moroz LA, Gilmore NJ. 1975; A rapid and sensitive ¹²⁵I-fibrin solidphase fibrinolytic assay for plasmin. Blood 46: 543-553
• 9 Nanninga LB, Guest MM. 1968; Activity-pH relationship and Michaelis constants during activation of profibrinolysin and during fibrinogenolysis. Thromb. Diath. Haemorrh 19: 492-498
• 10 Ratnoff OD. 1969; Some relationships among hemostasis, fibrinolytic phenomena, immunity, and the inflammatory response. Adv. Immunol 10: 145-227
• 11 Remmert LF, Cohen PP. 1949; Partial purification and properties of a proteolytic enzyme of human serum. J. Biol. Chem.. 181: 431-448
• 12 Sawyer WD, Fletcher AP, Alkjaersig N, Sherry S. 1969; Studies on the thrombolytic activity of human plasma. J. Clin. Invest 39: 426-434
• 13 Weber K, Osborn M. 1969; The reliability of molecular weight determination by dodecyl sulphate-polyacrylamide gel electrophoresis. J. Biol. Chem. 244: 4406-4412
• 14 Wiman B, Wallen P. 1977; The specific interaction between plasminogen and fibrin. A physiological role of the lysine binding site in plasminogen Thromb. Res. 10: 213-222