Steady state kinetic parameters for the thrombin-catalyzed conversion of human fibrinogen to fibrin.
Steady state kinetic parameters were evaluated for the hydrolytic release of fibrinopeptides A and B (FPA and FPB) from human fibrinogen by human thrombin at pH 7.4, 37 degrees C, and gamma/2 0.15. At low concentrations of fibrinogen (less than 0.4 microM), the release of FPA from A alpha-chains was first order with respect to both the concentration of fibrinogen A alpha-chains and thrombin. The second order rate constant yielded a value of 11.6 (+/- 0.3) X 10(6) M-1 S-1 for the specificity constant (kcat/Km) for this process. Values of 84 (+/- 4) S-1 and 7.2 (+/- 0.9) microM were evaluated for kcat and Km for the thrombin-catalyzed release of FPA from normal human fibrinogen. The amino acid replacement ArgA alpha 16 leads to His present in fibrinogen Petoskey was shown to result in a 160-fold decrease in the specificity constant for hydrolysis at A alpha 16 and concomitant release of FPA. A kinetic analysis for determination of the sequentiality of release of fibrinopeptides was presented. It indicated that at least 97% of FPB was released after FPA. The specificity constant for release of FPB from intact fibrinogen (if it occurs) was less than 3% of that for release of FPA and less than 10% of that for release of FPB from des-A fibrinogen. The specificity constant for the release of FPB from des-A fibrinogen was 4.2 (+/- 0.2) X 10(6) M-1 S-1. The polymerization inhibitors EDTA and Gly-Pro-Arg-Pro inhibited release of FPB but not FPA. These observations are consistent with the generally accepted view that the predominant pathway for the conversion of normal human fibrinogen to fibrin is one wherein FPA is released, des-A fibrinogen polymerizes, and then FPB is released.
D004492:Edetic Acid / Q000494:pharmacology; D005337:Fibrin / Q000378:metabolism*; D005340:Fibrinogen / Q000378:metabolism*; D005344:Fibrinopeptide A / Q000378:metabolism; D005345:Fibrinopeptide B / Q000378:metabolism; D006801:Humans; D007700:Kinetics; D008433:Mathematics; D009842:Oligopeptides / Q000494:pharmacology; D013917:Thrombin / Q000378:metabolism*
D009842:Oligopeptides; D005344:Fibrinopeptide A; D005345:Fibrinopeptide B; C034585:glycyl-prolyl-arginyl-proline; D005337:Fibrin; D005340:Fibrinogen; D004492:Edetic Acid; D013917:Thrombin