Bovine Thrombin Complexed with an Uncleavable Analog of Residues 7−19 of Fibrinogen Aα:  Geometry of the Catalytic Triad and Interactions of the P1‘, P2‘, and P3‘ Substrate Residues

The crystal structure of the noncovalent complex of bovine thrombin and a fibrinogen-Aα tridecapeptide substrate analog, G17ψ, in which the scissile bond amide nitrogen of Gly-17f has been replaced by a methylene carbon, has been determined at 2.3 Å resolution with an R factor of 17.1%. The geometry of the active site indicates that the crystal structure is a close model of the true Michaelis complex. The three independently determined thrombin/G17ψ complexes in the crystal asymmetric unit reveal novel interactions for the P2‘ and P3‘ residuesPro-18f and Arg-19f, respectivelyon the carboxyl-terminal side of the scissile bond and confirm previously observed interactions of the P1 (Arg-16f) through P10 (Asp-7f) positions on the amino-terminal side. The thrombin S2‘ binding site for Pro-18f, as observed in all three complexes, differs from that predicted by modeling studies and is notable for including two carbonyl oxygens of the thrombin main chain. Arg-19f occupies two binding sites on thrombin, S3‘A and S3‘B, which have dramatically different placements for the arginyl side chain and carboxyl terminus.