血栓性疾病是一类严重危害人类尤其是中老年人健康和生命的疾病，近年来其发病率和致死率逐年上升，因此开展与其相关的基础研究和新药开发十分迫切。人体维生素K循环中的限速酶hVKOR是开发新型抗凝药物和维生素K拮抗剂的解毒剂的靶点。临床上常用的抗凝药物华法林就是通过特异性靶向抑制hVKOR，达到抗凝血的效果，但其用药存在个体差异大，治疗窗口窄的问题。目前hVKOR在维生素K循环中的催化机制还不清楚，且华法林如何抑制hVKOR也存在争议。我们前期研究发现hVKOR在催化过程中与其底物KO形成了一种过渡态复合物，并初步构建了hVKOR催化KO的模型。本项目拟在此基础上，采用hVKOR活性实验、蛋白质谱、分子模拟和蛋白结构解析等方法系统地研究和阐明hVKOR的催化机制、华法林抑制hVKOR的机制和hVKOR活性中心活化的分子基础。本研究将为研发新型抗凝药物和维生素K拮抗剂的解毒剂提供理论指导。

Thrombotic disease is a kind of serious hazard disease to human health and life, especially in middle-aged an elderly people. Recently, the morbidity and mortality rate of thrombotic disease increased year by year, so it is urgent to carry out basic research and new drug development of thrombotic disease. Human Vitamin K epoxide reductase (hVKOR), a speed-limit enzyme in vitamin K cycle, is the target to develop new anticoagulants and antidotes of vitamin K antagonists. Warfarin, an usually used anticoagulant in clinical to inhibit blood coagulation, is such kind of drug to specifically target and inhibit hVKOR. However, There are problems of individual difference and narrow therapeutic window in use of warfarin. Currently, it is still unclear about the catalytic mechanism of hVKOR, and also controversial about the mechanism of warfarin inhibiting hVKOR. Our preliminary data found that hVKOR forms an intermediate complex with its substrate vitamin K epoxide (KO) during catalytic process, and we attempt to build a model of hVKOR catalyzing KO. On the above basis, this project will combine methods of hVKOR activity assay, protein mass spectrometry, molecular dynamics simulation, protein structure determination, et al, to systemically study catalytic mechanism of hVKOR, mechanism of warfarin inhibiting hVKOR, and molecular basis of hVKOR active site activating. So this study will provide important theoretical direction for the design of new specific anticoagulants and antidotes of vitamin K antagonists.