Structure-Function Relationship of CAR

描述(由申请人提供)：这项建议的长期目标是了解构成雄烷受体(CAR)的结构机制。CAR属于核受体超家族蛋白质。这些都是激素调节的转录因子，几乎指导着人类生理的方方面面。在没有配基的情况下，CAR具有结构性活性。已经发现了调节汽车活动的小分子。类固醇分子雄烷醇起反向激动剂的作用，抑制CAR的构成活性。另一种分子，TCPOBOP，将CAR从雄烷醇介导的抑制中拯救出来，并可以增强CAR的活性。我们假设，汽车结构采用了独特的，但微妙的修改，从规范的核受体结构折叠，导致这种独特的行为。我们假设CAR的三个状态，即非活性状态、雄烷醇结合状态、活性状态、载脂蛋白和TCPOBOP结合的高活性状态通过变构机制联系在一起。在具体目标1中，我们建议比较载脂蛋白Car和雄烯醇结合的Car的结构。在具体目标2中，我们将比较CAR：TCPOBOP与载脂蛋白和雄烷醇结合的CAR结构。在具体目标3中，我们将通过对这些突变体进行基于结构的点突变以及结合和活性分析来研究CAR变构的机制。 细胞色素P50是细胞色素P50家族中的一类酶，CAR被证明能够指导细胞色素P50基因产物的转录。这些酶负责外源物质的新陈代谢，如药物。CAR直接参与了止痛药、对乙酰氨基酚(商业术语：泰诺)等药品的代谢。了解CAR的分子机制在核受体生物学中具有重要意义，并在这些蛋白质的药理学中具有广泛的意义。由于参与了治疗药物的代谢，CAR本身就是设计小分子配体的一个有吸引力的靶点，这些小分子配体可以增强或减弱其转录活性。

DESCRIPTION (provided by applicant): The long-term goal of this proposal is to understand the structural mechanism of the constitutive androstane receptor (CAR). CAR belongs to the nuclear receptor superfamily of proteins. These are hormone-modulated transcription factors that direct almost every aspect of human physiology. CAR is constitutively active in the absence of ligand. Small molecules have been identified that modulate CAR activity. The steroidal molecule, androstanol, functions as an inverse agonist and inhibits the constitutive activity of CAR. Another molecule, TCPOBOP, rescues CAR from the androstanol-mediated repression and can enhance CAR activity. We hypothesize that the CAR structure employs unique, but subtle, modifications from the canonical nuclear receptor structural fold that results in this distinctive behavior. We hypothesize that the three states of CAR, the inactive, androstanol-bound, the active, apo, and the TCPOBOP-bound highly active states are linked by an allosteric mechanism. In Specific Aim 1 we propose to compare the structures of apo CAR with androstenol-bound CAR. In Specific Aim 2, we will compare the CAR:TCPOBOP to apo and androstanol-bound CAR structures. In Specific Aim 3, we will study the mechanism of CAR allosterism by performing structure-based point mutations and binding and activity assays on these mutants. CAR has been shown to direct the transcription of the CYP2 and CYP3 gene products, which belong to the cytochrome P50 class of enzymes. These enzymes are responsible for the metabolism of xenobiotics such as pharmaceutical drugs. CAR has been directly implicated in the metabolism of the analgesic, acetaminophen (commercial nomenclature: Tylenol) other pharmaceutical products. Understanding the CAR molecular mechanism is of great significance in nuclear receptor biology with broad implications in the pharmacology of these proteins. Because of its involvement in metabolism of therapeutics, CAR is itself an attractive target for the design of small molecule ligands that can both enhance or diminish its transcriptional activity.