蚋虫唾液蛋白Sibanin抑制宿主止血的机制研究

蚋虫是一种重要的吸血节肢动物，通过吸食宿主的血液获取营养进行产卵和繁殖，然而其血餐机制不明。我们从云南特有蚋虫——版纳绳蚋的唾液腺中纯化出一个结构新颖的蛋白，命名为Sibanin。它能够抑制由ADP诱导的血小板聚集；抑制由胶原诱导的血小板聚集；抑制FXa（活化的凝血因子X）的活性，显著延长血浆凝固时间。我们推断Sibanin通过抑制血小板聚集和抑制FXa活性的双重机制来对抗宿主的止血反应。本项目拟在前期工作基础上，明确Sibanin作用于血小板上的受体和相关蛋白，阐明其抑制血小板聚集的分子机制；明确其作用于FXa上的靶点，测定其在酶促反应中的动力学参数如Ki、Ka等，阐明其抑制FXa的作用机制和特性；明确其在小鼠和大鼠体内抑制血栓形成的作用，评估其可能引起的出血风险，阐明其在体内的抗血栓功效和副作用。本项目将为揭示版纳绳蚋的血餐机制提供直接的分子依据，并为抗血栓药物研发提供潜在候选分子。

The black fly is an important blood-sucking arthropod, and its blood-meal provides the nutrition for the egg-laying and reproduction. However, the mechanism of blood-meal is poorly understood. Recently, one protein named Sibanin was purified from the salivary glands of the black fly, simulium bannaense. Sibanin inhibited the platelet aggregation induced by ADP, and it also inhibited the platelet aggregation induced by collagen. Additionally, Sibanin inhibited the activity of activated coagulation factor X (FXa) and significantly prolonged the clotting time. We infer that Sibanin counteracts the hemostatic response of the host through the dual mechanism of inhibiting platelet aggregation and inhibiting the activity of FXa. Based on the preliminary work, we intend to investigate the receptors of platelets and related proteins that Sibanin acts on, clarifying its molecular mechanism for inhibiting platelet aggregation. We also will investigate the effect of Sibanin on the active site of FXa and measure its kinetic parameters in the enzymatic reaction such as inhibition constant (Ki), association constant(Ka), etc., clarifying its mechanism and characteristics of inhibiting FXa. In addition, we will investigate the effect of Sibanin in inhibiting thrombosis in mice and rats, and detect its possible hemorrhagic activity, clarifying its antithrombotic effects and side effects in vivo. This project will provide a direct molecular basis for revealing the blood-meal mechanism of simulium bannaense and provide a potential candidate molecule for the development of antithrombotic drugs.