Wei A, Alexander R S, Duke J, Ross H, Rosenfeld S A, Chang C H
Department of Chemical and Physical Sciences, The DuPont Pharmaceuticals Company, Wilmington DE 19880, USA.
J Mol Biol. 1998;283(1):147-54. doi: 10.1006/jmbi.1998.2069.
The structure of recombinant tick anticoagulant peptide (rTAP) complexed to bovine factor Xa at 3.0 A resolution reveals the structural basis for the specificity and the high affinity of rTAP. Three N-terminal residues, Tyr501, Asn502 and Arg503, play a critical role in the complex formation as suggested by earlier mutagenic studies and the ornithodorin-thrombin complex. Unexpectedly, the side-chain of Tyr501 is located in the S1 pocket, although factor Xa favors arginine as a P1 residue. Arg503 is located at the aryl binding pocket and forms a salt-bridge with Glu97 of factor Xa. The autolysis loop, which is disordered in the uninhibited factor Xa structure, is involved in the formation of the complex as a part of the secondary binding site. The C-terminal helix of rTAP interacts with factor Xa as a secondary binding determinant. The N-terminal residues of rTAP reorganize during the formation of the factor Xa-rTAP complex from the conformation found in the solution into an extended conformation. The presence of the secondary binding site confirms the proposed two-step kinetic mechanism based on the results of a mutagenesis study.
以3.0埃分辨率解析的重组蜱抗凝肽（rTAP）与牛凝血因子Xa的复合物结构揭示了rTAP特异性和高亲和力的结构基础。如早期诱变研究和鸟氨酸蛋白酶-凝血酶复合物所示，三个N端残基Tyr501、Asn502和Arg503在复合物形成中起关键作用。出乎意料的是，尽管凝血因子Xa倾向于精氨酸作为P1残基，但Tyr501的侧链位于S1口袋中。Arg503位于芳基结合口袋，与凝血因子Xa的Glu97形成盐桥。在未受抑制的凝血因子Xa结构中无序的自溶环作为二级结合位点的一部分参与复合物的形成。rTAP的C端螺旋作为二级结合决定因素与凝血因子Xa相互作用。在凝血因子Xa-rTAP复合物形成过程中，rTAP的N端残基从溶液中发现的构象重新组织成伸展构象。二级结合位点的存在证实了基于诱变研究结果提出的两步动力学机制。
