Stabilization of the E* form turns thrombin into an anticoagulant.
Previous studies have shown that deletion of nine residues in the autolysis loop of thrombin produces a mutant with an anticoagulant propensity of potential clinical relevance, but the molecular origin of the effect has remained unresolved. The x-ray crystal structure of this mutant solved in the free form at 1.55 A resolution reveals an inactive conformation that is practically identical (root mean square deviation of 0.154 A) to the recently identified E* form. The side chain of Trp(215) collapses into the active site by shifting > 10 A from its position in the active E form, and the oxyanion hole is disrupted by a flip of the Glu(192)-Gly(193) peptide bond. This finding confirms the existence of the inactive form E* in essentially the same incarnation as first identified in the structure of the thrombin mutant D102N. In addition, it demonstrates that the anticoagulant profile often caused by a mutation of the thrombin scaffold finds its likely molecular origin in the stabilization of the inactive E* form that is selectively shifted to the active E form upon thrombomodulin and protein C binding.
Bah
Alaji
A
Department of Biochemistry and Molecular Biophysics, Washington University School of Medicine, St. Louis, Missouri 63110, USA.
Carrell
Christopher J
CJ
Chen
Zhiwei
Z
Gandhi
Prafull S
PS
Di Cera
Enrico
E
D000925:Anticoagulants / Q000737:chemistry* / Q000378:metabolism*; D001120:Arginine / Q000031:analogs & derivatives; D001665:Binding Sites; D018360:Crystallography, X-Ray; D004789:Enzyme Activation; D006801:Humans; D008958:Models, Molecular; D016297:Mutagenesis, Site-Directed; D050505:Mutant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D010875:Pipecolic Acids / Q000378:metabolism; D011485:Protein Binding; D011486:Protein C / Q000378:metabolism; D011487:Protein Conformation; D013449:Sulfonamides; D013917:Thrombin / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D018180:Thrombomodulin / Q000378:metabolism
D000925:Anticoagulants; D050505:Mutant Proteins; D010875:Pipecolic Acids; D011486:Protein C; D013449:Sulfonamides; D018180:Thrombomodulin; D001120:Arginine; D013917:Thrombin; C031942:argatroban