Bovine thrombin complexed with an uncleavable analog of residues 7-19 of fibrinogen A alpha: 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a fibrinogen-A alpha tridecapeptide substrate analog, G17 psi, in which the scissile bond amide nitrogen of Gly-17f has been replaced by a methylene carbon, has been determined at 2.3 A resolution with an R factor of 17.1%.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/G17 psi 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.
Martin
P D
PD
Department of Biochemistry, Wayne State University, Detroit, Michigan 48201, USA.
Malkowski
M G
MG
DiMaio
J
J
Konishi
Y
Y
Ni
F
F
Edwards
B F
BF
D000818:Animals; D001665:Binding Sites; D002417:Cattle; D018360:Crystallography, X-Ray; D005340:Fibrinogen / Q000737:chemistry*; D006629:Hirudins / Q000031:analogs & derivatives / Q000737:chemistry; D006860:Hydrogen Bonding; D008958:Models, Molecular; D010446:Peptide Fragments / Q000737:chemistry*; D011487:Protein Conformation; D013917:Thrombin / Q000737:chemistry* / Q000378:metabolism
C101922:G17psi peptide; D006629:Hirudins; D010446:Peptide Fragments; C085066:hirutonin-2; D005340:Fibrinogen; D013917:Thrombin