STRUCTURE/FUNCTION STUDIES OF ANTIHEMOSTATIC LIPOCALINS

食血昆虫的唾液中含有多种止血物质。 帮助获得不间断的血液流向 口罩。这些药物包括血管扩张剂、血小板聚集抑制剂、 和抗凝血剂。这些物质中的大多数都是蛋白质类物质，而且 他们的目标已经确定了一些。然而，分子 他们的行动机制和负责的结构特征 他们的活动在很大程度上是未知的。这项建议侧重于两种类型 三甲虫Rhodnius唾液中抗止血药物的含量 Prolixus，抗凝剂Nitphorin 2和三种已知的蛋白质 唾液抗血小板脂蛋白(SAPLs)。亚硝基肾上腺素是一组 四种转运一氧化氮和结合一氧化氮的多功能血红蛋白 组胺。Nitrophorin 2是该集团中唯一拥有 也有抗凝血活性。SAPLs 1-3抑制胶原蛋白介导 血小板聚集和凝血酶活性。令人惊讶的是，这两个 硝基酚和SAPL属于Lipocalin蛋白家族，尽管 他们完全不同的活动。在这些测试中检验的假设 研究表明，硝基苯酚2的抗凝血活性和抗凝血酶活性。 SAPLs 1-3的抗血小板活性是由于这些物质相互作用所致 具有特定蛋白质靶点或与必需物质结合的分子 配基。这项研究的具体目的是分析这些机制 和硝基苯酚2和SAPLs 1-3活性的结构决定因素 使用功能分析、X射线结晶学和定点定位 诱变。

The saliva of blood-feeding insects contains a variety of antihemostatic agents that aid in obtaining an uninterrupted flow of blood to the mouthparts. These include vasodilators, platelet aggregation inhibitors, and anticoagulants. Most of these substances are proteinaceous, and a number of their targets have been identified. However, the molecular mechanisms for their actions and the structural features responsible for their activity are largely unknown. This proposal focuses on two types of antihemostatic agents from the saliva of the triatomine bug Rhodnius prolixus, the anticoagulant nitrophorin 2 and three proteins known as salivary antiplatelet lipocalins (SAPLs). Nitrophorins are a group of four multifunctional hemeproteins which transport nitric oxide and bind histamine. Nitrophorin 2 is the only member of the group possessing anticoagulant activity as well. SAPLs 1-3 inhibit collagen-mediated platelet aggregation and thrombin activity. Surprisingly, both the nitrophorins and SAPLs belong to the lipocalin protein family despite their completely different activities. The hypotheses tested in these studies are that the anticoagulant activity of nitrophorin 2 and the antiplatelet activity of SAPLs 1-3 are due to interaction of these molecules with specific proteinaceous targets or binding of essential ligands. The specific aims of the study are to analyze the mechanisms and structural determinants of activity of nitrophorin 2 and SAPLs 1-3 using functional assays, X-ray crystallography, and site-directed mutagenesis.