Recombinant hirudin: kinetic mechanism for the inhibition of human thrombin
Stone, Stuart R. ; Hofsteenge, Jan
In: Protein Engineering, Design and Selection, 1991, vol. 4, no. 3, p. 295-300
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- Recombinant hirudin variant-2(Lys47 ), was found to be a competitive inhibitor of human α-thrombin with respect to peptidyl p-Miitroanilide substrates. These results contrast with those of Degryse and coworkers that suggest that recombinant hirudin variant-2(Lys47) inhibited thrombin by a non-competitive mechanism [Degryse et al. (1989) Protein Engng, 2, 459-465], γ-Thrombin, which can arise from α-thrombin by autolysis, was shown to have an affinity for recombinant hirudin variant-2(Lys47) that was four orders of magnitude lower than that of α-thrombin. It was demonstrated that the apparent noncompetitive mechanism observed previously was probably caused by a contamination of the thrombin preparation by γ-thrombin. Comparison of the inhibition of α-thrombin by recombinant hirudins variant-2(Lys47) and variant-1, which differ from one another in eight out of 65 amino acids, indicated that the two variants have essentially the same kinetic parameters