Wang Stephanie X, Hur Eugene, Sousa Carolyn A, Brinen Linda, Slivka Eric J, Fletterick Robert J
Graduate Program in Chemistry and Chemical Biology, University of California, San Francisco, California 94143, USA.
Biochemistry. 2003 Jul 8;42(26):7959-66. doi: 10.1021/bi027320a.
The serine protease factor Xa (FXa) is inhibited by ecotin with picomolar affinity. The structure of the tetrameric complex of ecotin variant M84R (M84R) with FXa has been determined to 2.8 A. Substrate directed induced fit of the binding interactions at the S2 and S4 pockets modulates the discrimination of the protease. Specifically, the Tyr at position 99 of FXa changes its conformation with respect to incoming ligand, changing the size of the S2 and S4 pockets. The role of residue 192 in substrate and inhibitor recognition is also examined. Gln 192 from FXa forms a hydrogen bond with the P2 carbonyl group of ecotin. This confirms previous biochemical and structural analyses on thrombin and activated protein C, which suggested that residue 192 may play a more general role in mediating the interactions between coagulation proteases and their inhibitors. The structure of ecotin M84R-FXa (M84R-FXa) also reveals the structure of the Gla domain in the presence of Mg(2+). The first 11 residues of the domain assume a novel conformation and likely represent an intermediate folding state of the domain.
丝氨酸蛋白酶因子Xa（FXa）可被艾考汀以皮摩尔亲和力抑制。已确定艾考汀变体M84R（M84R）与FXa的四聚体复合物结构，分辨率达2.8埃。底物导向的结合相互作用在S2和S4口袋处的诱导契合调节了蛋白酶的识别。具体而言，FXa第99位的酪氨酸相对于进入的配体改变其构象，从而改变S2和S4口袋的大小。还研究了192位残基在底物和抑制剂识别中的作用。FXa的谷氨酰胺192与艾考汀的P2羰基形成氢键。这证实了先前对凝血酶和活化蛋白C的生化和结构分析，表明192位残基可能在介导凝血蛋白酶与其抑制剂之间的相互作用中发挥更普遍的作用。艾考汀M84R - FXa（M84R - FXa）的结构还揭示了在Mg(2+)存在下Gla结构域的结构。该结构域的前11个残基呈现出一种新的构象，可能代表该结构域的一种中间折叠状态。
