Bovine alpha- and beta-thrombin. Reduced fibrinogen-clotting activity of beta-thrombin is not a consequence of reduced affinity for fibrinogen.
beta-Thrombin, a product of the limited proteolysis of alpha-thrombin, is characterized by greatly reduced fibrinogen-clotting activity as compared to alpha-thrombin but with unchanged activity toward ester substrates. The present study was designed to elucidate the basis for the changes in the catalytic activity resulting from the conversion of bovine alpha-thrombin to bovine beta-thrombin. Fibrinogen was utilized as a competitive inhibitor in the hydrolysis of a peptide nitroanilide substrate by bovine alpha- and beta-thrombin. The Ki values obtained for fibrinogen in these experiments were similar for alpha- and beta-thrombin (about 10 microM). Similar values for Ki were obtained when fibrinogen was used to inhibit the inactivation of bovine alpha- and beta-thrombin by diisopropylphosphorofluoridate. These experiments suggested that the conversion of bovine alpha- to beta-thrombin does not affect the fibrinogen-binding site on thrombin. Differences in the reactivity of functional groups at the active site were then explored. beta-Thrombin was observed to undergo modification at the active site histidine at a slower rate than that of alpha-thrombin when reacted with either tosyllysyl chloromethyl ketone or diethyl pyrocarbonate. It is suggested that the difference in the fibrinogen-clotting activity of these two forms of thrombin can result from changes in the reactivity of the active site histidine residue.
D000345:Affinity Labels / Q000378:metabolism; D000818:Animals; D002417:Cattle; D004047:Diethyl Pyrocarbonate / Q000494:pharmacology; D004151:Dipeptides / Q000378:metabolism; D005340:Fibrinogen / Q000378:metabolism*; D007531:Isoflurophate / Q000494:pharmacology; D007700:Kinetics; D046911:Macromolecular Substances; D008433:Mathematics; D013917:Thrombin / Q000378:metabolism*; D014107:Tosyllysine Chloromethyl Ketone / Q000378:metabolism
D000345:Affinity Labels; D004151:Dipeptides; D046911:Macromolecular Substances; D007531:Isoflurophate; D014107:Tosyllysine Chloromethyl Ketone; C021129:S 2238; D005340:Fibrinogen; D013917:Thrombin; D004047:Diethyl Pyrocarbonate