Molecular Design and Characterization of an -Thrombin Inhibitor Containing a Novel P1 Moiety
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- 作者：J. A. Malikayil ; Joseph P. Burkhart ; Herman A. Schreuder ; Robert J. Broersma ; Jr. ; Chantal Tardif ; Louis W. Kutcher ; III ; Shujaath Mehdi ; Gerald L. Schatzman ; Bernhard Neises ; and Norton P. Peet
- 刊名：Biochemistry
- 出版年：1997
- 出版时间：February 4, 1997
- 年：1997
- 卷：36
- 期：5
- 页码：1034 - 1040
- 全文大小：170K
- 年卷期：v.36,no.5(February 4, 1997)
- ISSN：1520-4995
文摘
An inhibitor of 
-thrombin was designed on the basis of theX-ray crystal structures of thrombinand trypsin. The design strategy employed the geometric andelectrostatic differences between thespecificity pockets of the two enzymes. These differences arisedue to the replacement of Ser 190 intrypsin by Ala 190 in thrombin. The new inhibitor contained atryptophan side chain instead of thearginine side chain that is present in the prototypical thrombininhibitors. This inhibitor had a Kivalueof 0.25 
M, displayed more than 400-fold specificity for thrombinover trypsin, and doubled the ratplasma APTT at a concentration of 44.9 M. The X-ray crystalstructure of the inhibitor/-thrombincomplex was determined. This represents the first reportedthree-dimensional structure of a thrombin/inhibitor complex where the specificity pocket of the enzyme is occupiedby a chemical moiety otherthan a guanidino or an amidino group. As was predicted by themolecular model, the tryptophan sidechain docks into the specificity pocket of the enzyme. Thisfinding is in contrast with the indole bindingregion of thrombin reported earlier [Berliner, L. J., & Shen, Y. Y. L.(1977) Biochemistry 16, 4622-4626]. The lower binding affinity of the new inhibitor fortrypsin, compared to that for thrombin, appearsto be due to (i) the extra energy required to deform the smallerspecificity pocket of trypsin to accommodatethe bulky indole group and (ii) the favorable electrostaticinteractions of the indole group with the morehydrophobic specificity pocket of thrombin. The neutral indolegroup may be of pharmacologicalsignificance because the severe hypotension and respiratory distressobserved following the administrationof some thrombin inhibitors have been linked to the positively chargedguanidino or amidino functionalities.
-thrombin was designed on the basis of theX-ray crystal structures of thrombinand trypsin. The design strategy employed the geometric andelectrostatic differences between thespecificity pockets of the two enzymes. These differences arisedue to the replacement of Ser 190 intrypsin by Ala 190 in thrombin. The new inhibitor contained atryptophan side chain instead of thearginine side chain that is present in the prototypical thrombininhibitors. This inhibitor had a Kivalueof 0.25 M, displayed more than 400-fold specificity for thrombinover trypsin, and doubled the ratplasma APTT at a concentration of 44.9 M. The X-ray crystalstructure of the inhibitor/-thrombincomplex was determined. This represents the first reportedthree-dimensional structure of a thrombin/inhibitor complex where the specificity pocket of the enzyme is occupiedby a chemical moiety otherthan a guanidino or an amidino group. As was predicted by themolecular model, the tryptophan sidechain docks into the specificity pocket of the enzyme. Thisfinding is in contrast with the indole bindingregion of thrombin reported earlier [Berliner, L. J., & Shen, Y. Y. L.(1977) Biochemistry 16, 4622-4626]. The lower binding affinity of the new inhibitor fortrypsin, compared to that for thrombin, appearsto be due to (i) the extra energy required to deform the smallerspecificity pocket of trypsin to accommodatethe bulky indole group and (ii) the favorable electrostaticinteractions of the indole group with the morehydrophobic specificity pocket of thrombin. The neutral indolegroup may be of pharmacologicalsignificance because the severe hypotension and respiratory distressobserved following the administrationof some thrombin inhibitors have been linked to the positively chargedguanidino or amidino functionalities.