Cholesterol Oxidase Loop Residues and Catalysis

描述（由申请人提供）：胆固醇氧化酶是一种黄酮类酶，可催化胆固醇氧化和异构化为胆固醇-4-烯-3- 1。胆固醇氧化酶在商业上用于测定血清胆固醇浓度，具有杀虫作用，并用于研究膜结构。最近的证据表明，胆固醇氧化酶可能参与一些微生物的发病机制，如马红球菌和结核分枝杆菌。提出的研究将提供一个精确的分子模型的脂质需求的活动和途径黄素氧化。这些模型对于开发胆固醇氧化酶的商业化产品、研究细胞膜、监测脂质层、研究胆固醇氧化酶在细菌发病机制中的作用以及开发抗菌抑制剂具有重要意义。将要测试的具体假设是：1)胆固醇氧化酶可以区分同一膜中不同组成的含胆固醇脂质结构域，即液体无序结构域和液体有序结构域（筏）；2)酶在催化过程中采用大界面与脂质双分子层结合；3)在溶剂和黄素的活性位点之间有一个门控通道控制氧气的进入。

DESCRIPTION (provided by applicant): Cholesterol oxidase is a flavoenzyme that catalyzes the oxidation and isomerization of cholesterol to cholest-4-en-3-one. Cholesterol oxidase is used commercially for the determination of serum cholesterol concentrations, is insecticidal, and is used to study membrane structure. Recent evidence suggests that cholesterol oxidase may be involved in the pathogenesis of some microorganisms such as Rhodococcus equi and Mycobacterium tuberculosis. The studies proposed will provide a precise molecular model of the lipid requirements for activity and of the pathway to flavin oxidation. These models are important for the development of cholesterol oxidase as a commercial product, as a useful and reliable tool in the study of cellular membranes for monitoring lipid rafts, for investigating the role of cholesterol oxidase in bacterial pathogenesis, and for developing anti-bacterial inhibitors. The specific hypotheses that will be tested are that 1) cholesterol oxidase can distinguish between cholesterol-containing lipid domains of different composition in the same membrane, i.e., between liquid-disordered domains and liquid-ordered domains (rafts); 2) the enzyme employs a large interface to associate with the lipid bilayer during catalysis; and 3) a gated tunnel between the solvent and the flavin at the active site controls oxygen access.