STUCTURE FUNCTION AND REGULATION OF NADPH CYTOCHROME P450 OXIDOREDUCTASE

NADPH细胞色素P450氧化还原酶的结构、功能及调控

• 批准号: 6300172
• 负责人: CHARLES B KASPER
• 金额: $ 23.98万
• 依托单位: UNIVERSITY OF WISCONSIN-MADISON
• 依托单位国家: 美国
• 财政年份: 2000
• 资助国家: 美国
• 起止时间: 2000-02-11 至 2001-01-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6300172
• 关键词: NAD(P)H oxidoreductase; active sites; chemical carcinogenesis; cytochrome P450; enzyme induction /repression; enzyme mechanism; enzyme structure; enzyme substrate complex; gel mobility shift assay; gene expression; gene induction /repression; immunocytochemistry; in situ hybridization; laboratory mouse; laboratory rat; molecular oncology; nucleic acid sequence; protein structure function

In a biochemical sense, the mixed-function oxidase system represents the first line of defence between man and his environment. This system is responsible for the early metabolic steps in they detoxification of xenobiotics as well as the metabolic activation of certain chemical carcinogens. Hence, the response of the cell to foreign compounds and the eventual fate of the cell depends upon the activity level of these key enzymes. This proposal is an extension of our ongoing studies dealing with the structure. function, and regulation of NADPH-cytochrome P450 oxidoreductase (P450R) and the cytochrome 3A family. Four major areas will be addressed during the next granting period. The first will examine those structural features of the reductase molecule that are responsible for maintaining optimal alignment and integrity of the flavin domains. Three specific regions will be analyzed. These are the interconnecting flavin domain, the carboxyl terminal region that is important for the structural and functional character of the FAD/NADPH binding domains and is the port of entry for electrons from NADPH, and the hinge region close to the FMN domain. The second area will focus on identify the recognition site(s) on the surface of P450R responsible for the selective binding of cytochrome P450 and other protein substrates such as cytochrome b5 and heme oxygenase. Third, the regulation of the P450R and CYP3A23 genes will be investigated in order to better understand the factors controlling the cellular levels of these enzymes. Fourth, the biological role of P450R will be evaluated by studying the developmental and tissue-specific expression using in situ hybridization and immunohistochemistry along with the creation of a mouse strain with a null allele at the P450R locus.

从生化意义上讲，混合功能氧化酶系统代表 人类与其环境之间的第一道防线。这个系统是 负责解毒的早期代谢步骤 外源物质以及某些化学物质的代谢激活 致癌物质。因此，细胞对外来化合物的反应和 细胞的最终命运取决于这些关键的活性水平 酶。 该提案是我们正在进行的研究的延伸，涉及 结构。 NADPH-细胞色素 P450 的功能和调节 氧化还原酶 (P450R) 和细胞色素 3A 家族。四大领域将 并在下一个授权期内予以解决。第一个将检查那些 负责还原酶分子的结构特征 保持黄素结构域的最佳排列和完整性。三 具体地区要具体分析。这些是互连黄素 结构域，对结构很重要的羧基末端区域 FAD/NADPH 结合域的功能特征和端口 来自 NADPH 的电子进入，以及靠近 FMN 的铰链区域 领域。第二个领域将侧重于识别识别位点 P450R 表面负责细胞色素的选择性结合 P450 和其他蛋白质底物，例如细胞色素 b5 和血红素 加氧酶。第三，P450R和CYP3A23基因的调控将被 进行调查，以便更好地了解控制因素 这些酶的细胞水平。四、P450R的生物学作用 将通过研究发育和组织特异性来评估 使用原位杂交和免疫组织化学进行表达 创建 P450R 基因座具有无效等位基因的小鼠品系。