Crystal structures of thrombin with thiazole-containing inhibitors: probes of the S1' binding site.
Structures of the blood clotting enzyme thrombin complexed with hirugen and two active site inhibitors, RWJ-50353 10080(N-methyl-D-phenylalanyl-N-[5-[(aminoiminomethyl)amino]-1- [[(2-benzothiazolyl)carbonyl]butyl]-L-prolinamide trifluoroacetate hydrate) and RWJ-50215 (N-[4-(aminoiminomethyl)amino-1-[2- (thiazol-2-ylcarbonylethyl)piperidin- 1-ylcarbonyl]butyl]-5-(dimethylamino)naphthalenesulfonamide trifluoroacetate hydrate), were determined by x-ray crystallography. The refinements converged at R values of 0.158 in the 7.0-2.3-A range for RWJ-50353 and 0.155 in the 7.0-1.8-A range for RWJ-50215. Interactions between the protein and the thiazole rings of the two inhibitors provide new valuable information about the S1' binding site of thrombin. The RWJ-50353 inhibitor consists of an S1'-binding benzothiazole group linked to the D-Phe-Pro-Arg chloromethyl ketone motif. Interactions with the S1-S3 sites are similar to the D-phenylalanyl-prolyl-arginyl chloromethylketone structure. In RWJ-50215, a S1'-binding 2-ketothiazole group was added to the thrombin inhibitor-like framework of dansylarginine N-(3-ethyl-1,5-pentanediyl)amide. The geometry at the S1-S3 sites here is also similar to that of the parent compound. The benzothiazole and 2-ketothiazole groups bind in a cavity surrounded by His57, Tyr60A, Trp60D, and Lys60F. This location of the S1' binding site is consistent with previous structures of thrombin complexes with hirulog-3, CVS-995, and hirutonin-2 and -6. The ring nitrogen of the RWJ-50353 benzothiazole forms a hydrogen bond with His57, and Lys60F reorients because of close contacts. The oxygen and nitrogen of the ketothiazole of RWJ-50215 hydrogen bond with the NZ atom of Lys60F.
Department of Chemistry, Michigan State University, East Lansing 48824, USA.
J H
JH
Matthews
R
R
Krishnan
M J
MJ
Costanzo
B E
BE
Maryanoff
A
A
Tulinsky
D001665:Binding Sites; D018360:Crystallography, X-Ray; D003619:Dansyl Compounds / Q000737:chemistry; D004791:Enzyme Inhibitors / Q000737:chemistry*; D006146:Guanidines / Q000737:chemistry*; D006629:Hirudins / Q000031:analogs & derivatives / Q000737:chemistry; D006801:Humans; D010446:Peptide Fragments / Q000737:chemistry; D011487:Protein Conformation; D013844:Thiazoles / Q000737:chemistry*; D013917:Thrombin / Q000037:antagonists & inhibitors / Q000648:ultrastructure*
D003619:Dansyl Compounds; D004791:Enzyme Inhibitors; D006146:Guanidines; D006629:Hirudins; D010446:Peptide Fragments; C104134:RWJ 50215; C100912:RWJ 50353; D013844:Thiazoles; C062510:hirugen; D013917:Thrombin