Structure-Function of Insect Odorant-Binding Proteins and a Bacterial Cytochrome P450: Discovery of Mechanisms and Applications

昆虫气味结合蛋白和细菌细胞色素 P450 的结构功能：机制的发现和应用

• 批准号: RGPIN-2020-05297
• 负责人: Plettner, Erika
• 金额: $ 2.62万
• 依托单位: Simon Fraser University
• 依托单位国家: 加拿大
• 项目类别: Discovery Grants Program - Individual
• 财政年份: 2020
• 资助国家: 加拿大
• 起止时间: 2020-01-01 至 2021-12-31
• 项目状态: 已结题
• 来源: https://www.nserc-crsng.gc.ca/ase-oro/Details-Detailles_eng.asp?id=716161
• 关键词: Structure; Function; Insect; Odorant; Binding

We propose to study two types of protein that interact with hydrophobic compounds: odorant binding proteins (OBPs) from insects and a cytochrome P450 from a soil bacterium. Both themes address problems of insect control in agriculture and forestry. OBPs are important insect-specific proteins, found in olfactory or pheromone-production systems. These proteins bind hydrophobic odorants, e.g. pheromones, compounds that are used for communication between members of the same species. The OBPs serve important functions in controlling the rate at which pheromones are detected or released. Here we propose to study OBPs from the gypsy moth, an economically important pest of trees, and from the honey bee, an economically important beneficial insect. In both cases, the OBPs of interest bind hydrophobic compounds: the gypsy moth pheromone ((+)-disparlure) is a hydrocarbon epoxide that is released by the females to attract males, whereas the honey bee pheromone (ethyl oleate) is a fatty acid ester that is produced by old worker bees that forage, to delay the transition of younger nurse bees to foraging activities. Transitioning nurse bees engage in hygienic behaviors, which make bees more resilient towards parasites and diseases. In both cases, we propose to study the structure of the OBP and its interactions with the pheromone, as well as interactions with the cognate odorant receptor in the olfactory system. In bees, we also propose to study how the OBP contributes to the transport of ethyl oleate from the stomach of the bee, where it forms, to the cuticle, where it is released. Cytochromes P450 are pigmented proteins that utilize dioxygen (O2) to oxidize hydrocarbons. P450cam catalyzes the oxidation of (+)-camphor under well-oxygenated conditions, but the reduction of camphor to borneol when the enzyme has been active in camphor oxidation, and O2 is depleted. We have found that formation of borneol affects the chemotactic behavior of the bacteria that contain P450cam, which hints at an ecological role of borneol formation when O2 is low. We seek to understand the mechanism of this camphor reduction, to understand its function. We have also discovered mutants of P450cam that dechlorinate the insecticide endosulfan, and related polychlorinated persistent pollutants. In order to develop mutants that dechlorinate these pollutants, we propose to study the mechanism of this process and optimize the mutants. Understanding the interactions of OBPs with pheromones will help us develop compounds that intervene in pheromone reception and alter insect behavior or inform the breeding of disease resistant bees. This work will be of interest to forestry and apiculture. Cytochromes P450 are potential biocatalysts for degradation of pollutants or production of new compounds. Work on polychlorinated pollutant degradation will be of value to the bioremediation community. Work on reactions of P450 will be valuable to the field of biocatalysis.

我们建议研究两种与疏水化合物相互作用的蛋白质：昆虫的气味结合蛋白(OBP)和土壤细菌的细胞色素P450。这两个主题都涉及农业和林业中的虫害控制问题。 OBP是重要的昆虫特有蛋白质，存在于嗅觉或信息素产生系统中。这些蛋白质结合了疏水性气味物质，例如信息素，这些化合物用于同一物种成员之间的交流。OBP在控制信息素被检测或释放的速度方面起着重要作用。在这里，我们打算研究吉普赛飞蛾和蜜蜂的OBP，吉普赛飞蛾是一种重要的经济害虫，蜜蜂是一种重要的经济有益昆虫。在这两种情况下，感兴趣的OBP结合疏水化合物：吉普赛飞蛾信息素((+)-dislure)是一种碳氢环氧化物，由雌性蜜蜂释放以吸引雄性，而蜜蜂信息素(油酸乙酯)是一种脂肪酸酯，由觅食的老工蜂产生，以延缓年轻哺乳蜜蜂到觅食活动的过渡。过渡的哺乳蜜蜂从事卫生行为，这使蜜蜂对寄生虫和疾病更具弹性。在这两种情况下，我们建议研究OBP的结构及其与信息素的相互作用，以及与嗅觉系统中同源气味感受器的相互作用。在蜜蜂中，我们还建议研究OBP如何促进油酸乙酯从蜜蜂的胃中运输到角质层，油酸乙酯在那里形成，在那里释放。 细胞色素P450是利用氧气(O2)氧化碳氢化合物的有色蛋白质。P450cam在氧气充足的条件下催化(+)-樟脑的氧化，但当该酶在樟脑的氧化中保持活性时，樟脑被还原为冰片，氧被耗尽。我们发现，冰片的形成影响了含有P450钙调素的细菌的趋化行为，这暗示了当氧气较低时，冰片的形成具有生态作用。我们试图了解这种樟脑还原的机制，以了解其功能。我们还发现了P450cam的突变体，它可以脱氯杀虫剂硫丹，以及相关的多氯持久性污染物。为了开发脱氯突变体，我们建议研究这一过程的机理，并对突变体进行优化。 了解OBP与信息素的相互作用将有助于我们开发干预信息素接收、改变昆虫行为或为培育抗病蜜蜂提供信息的化合物。这项工作对林业和养蜂业都有意义。细胞色素P450是降解污染物或产生新化合物的潜在生物催化剂。多氯污染物降解方面的工作将对生物修复社区有价值。对P450反应的研究将对生物催化领域具有重要的意义。