Characterization of the catalytic defect in the dysthrombin, Thrombin Quick.
The dysthrombin, Thrombin Quick, is chromatographically separable into two components designated Thrombin Quick I and Thrombin Quick II. Thrombin Quick II lacks observable catalytic activity toward thrombin substrates. The steady-state kinetics of hydrolysis of benzoylarginine ethyl ester and Tos-Gly-Pro-Arg-p-nitroanilide by Thrombin Quick I are equivalent to those of thrombin. These results, in addition to binding studies with the active site titrant N2-(5-dimethylaminonaphthalene-1-sulfonyl)arginine N-(3-ethyl-1,5-pentanediyl)amide, indicate that binding interactions at the catalytic site of Thrombin Quick I are unaltered. Thrombin Quick I is inhibited by anti-thrombin III at the same rate as thrombin. Steady-state kinetic parameters for the release of fibrinopeptide A indicate defects in both kcat and Km for Thrombin Quick I with kcat/Km equal to 0.012 of the value for thrombin, corresponding to the relative fibrinogen clotting activity of 0.013. The results are interpreted as indicating a defect in Thrombin Quick I at a binding site, external to the catalytic site, which is essential for determining specificity toward fibrinogen. The defect in kcat may result secondarily from small perturbations in the steric relationship of the catalytic triad residues. The rate of hydrolysis by Thrombin Quick I of the protein substrates bovine prothrombin and bovine protein C (in the absence of cofactors) is about one-third of that observed for thrombin, indicating that hydrolysis of these substrates by thrombin involves different specificity determinants than does the hydrolysis of fibrinogen.
D000990:Antithrombin III / Q000494:pharmacology; D001665:Binding Sites; D005340:Fibrinogen; D006801:Humans; D006868:Hydrolysis; D007700:Kinetics; D013379:Substrate Specificity; D013917:Thrombin / Q000037:antagonists & inhibitors / Q000302:isolation & purification / Q000378:metabolism*
D000990:Antithrombin III; D005340:Fibrinogen; D013917:Thrombin; C027648:thrombin Quick