WHY DO SUBSTRATES REQUIRE CYTOCHROME B5 FOR OXIDATION BY CYTOCHROME P450

为什么底物需要细胞色素 B5 才能被细胞色素 P450 氧化

• 批准号: 6280130
• 负责人: LUCY A WASKELL
• 金额: $ 0.11万
• 依托单位: UNIVERSITY OF CALIFORNIA, SAN FRANCISCO
• 依托单位国家: 美国
• 财政年份: 1998
• 资助国家: 美国
• 起止时间: 1998-07-01 至 1999-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6280130
• 关键词: biomedical resource; hematology; respiratory system

The overall long-term goal of our laboratory is to understand the molecular basis of the effect of cytochrome b5 (cyt b5) on the metabolism of certain substrates by cytochrome P-450 (cyt P-450) and to determine the physiological significance of this reaction. In particular, we are interested in elucidating the molecular basis of the marked stimulatory effect of cyt b5 on the metabolism of the volatile anesthetic, methoxyflurane, by cyt P-450 LM2 (2B4). These studies may eventually lead to the delineation of the etiology and pathophysiology of the postoperative hepatotoxicity attributed to the volatile anesthetics and to the development of safer anesthetics. These studies should also provide important information about the mechanism by which cyt P450 oxidizes its numerous endogenous and xenobiotic substrates. This knowledge should prove valuable in facilitating the design of clinically useful inhibitors of cytochrome P450 for therapeutic applications in fungal diseases, hyperaldosteronism, prostate cancer, benign prostatic hypertrophy and breast cancer. The more immediate short-term goal of this proposal is to use the 23,000 dalton cyt b5-cytochrome c (cyt c) complex as a model for the larger (72,000 dalton) hydrophobic cyt b5-cyt P450 complex and to determine the structure of the model complex using high resolution NMR. The cytochrome b5-cytochrome c complex is a relevant model for the larger hydrophobic complex because cytochrome b5 uses approximately the same site to interact with both cytochrome c and cytochrome P450. Knowledge of the structure of the cyt b5-cyt c complex will provide insights, into 1) the mechanism, 2) pathway of electron transfer, between cyt b5 and its redox partners, and 3) macromolecular recognition and protein dynamics. The appropriate three- and four-dimensional NMR experiments will be conducted on a 500 and 600 MHz NMR spectrometer with the available uniformly 13C/15N labeled bovine cyt b5 in the presence and absence of cyt c. Completion of the proposed NMR experiments should provide us with important structural details about how certain amino acids of cyt b5 facilitate electron transfer with its redox partners, cyt P-450, cyt b5 reductase and cyt c. The computing, graphics and programming facilities at the UCSF Computer Laboratory will be used to construct three-dimensional molecular models based on results from our NMR studies, to display and examine these models, and to calculate the dynamics and interactions of the components in the models; these results should lead to a greater understanding of the structural basis for the requirement for cyt b5 for the metabolism of some substrates by cyt P-450.

我们实验室的总体长期目标是了解 细胞色素 b5 (cyt b5) 对细胞色素 B5 影响的分子基础 细胞色素 P-450 (cyt P-450) 对某些底物的代谢和 以确定该反应的生理意义。 在 特别是，我们有兴趣阐明其分子基础 细胞色素b5对细胞代谢有显着的促进作用 挥发性麻醉剂甲氧氟烷，由 cyt P-450 LM2 (2B4) 提供。 这些 研究最终可能导致病因学的描述和 术后肝毒性的病理生理学 挥发性麻醉剂和开发更安全的麻醉剂。 这些研究还应该提供有关 cyt P450 氧化其众多内源性和 外源性底物。 这些知识应该被证明是有价值的 促进临床上有用的细胞色素抑制剂的设计 P450 用于真菌疾病的治疗应用， 醛固酮增多症、前列腺癌、良性前列腺肥大等 乳腺癌。 该提案更直接的短期目标是 使用 23,000 道尔顿的细胞色素 b5-细胞色素 c (cyt c) 复合物作为 较大（72,000 道尔顿）疏水性细胞色素 b5-细胞色素 P450 的模型 复杂并使用高确定模型复杂的结构 分辨率核磁共振。 细胞色素 b5-细胞色素 c 复合物是一个相关的 较大疏水复合物的模型，因为细胞色素 b5 使用 与细胞色素 c 和细胞色素 c 相互作用的位点大致相同 细胞色素P450。 了解 cyt b5-cyt c 的结构 复合体将提供对 1) 机制、2) 途径的见解 cyt b5 及其氧化还原伙伴之间的电子转移，以及 3) 大分子识别和蛋白质动力学。 适当的 三维和四维核磁共振实验将在 500 和 600 兆赫 核磁共振波谱仪可提供统一的 13C/15N 标记 存在和不存在 cyt c 时的牛 cyt b5。完成 拟议的核磁共振实验应该为我们提供重要的结构 有关 cyt b5 的某些氨基酸如何促进电子的详细信息 与其氧化还原伙伴、cyt P-450、cyt b5 还原酶和 cyt 一起转移 c. UCSF 的计算、图形和编程设施 计算机实验室将用于构建三维 基于我们 NMR 研究结果的分子模型，用于显示和 检查这些模型，并计算动力学和相互作用 模型中的组件；这些结果应该会导致 更好地理解需求的结构基础 cyt b5 通过 cyt P-450 代谢一些底物。