Structural Basis of Cytochrome P450 Activity

描述（由申请人提供）：外源代谢细胞色素P450 （P450）酶在药物代谢、毒素和致癌物原活化中起关键作用。由于每个P450都可以与广泛的底物和抑制剂相互作用，因此了解和预测外来化合物如何与这些酶结合并被这些酶代谢在毒理学，化学预防和多药学方面具有重要的实用价值。基于现有的人类细胞色素P450 2A和2E1酶的生物化学和结构生物学知识，本提案的目标是扩展，测试和应用我们对人类2A和2E酶结构与其配体选择性之间独特关系的理解。重要的是能够了解每一种酶的生理和药物代谢作用，以便操纵它们对人类健康产生积极影响。与这一建议相关的是，呼吸通路中的2A13激活烟草烟雾中两种最常见的致癌化合物之一，产生烷基化DNA的代谢物，并可引发肺癌。选择性抑制2A13将显著减少这一过程。具体来说，我们建议1)用生理相关和标记化合物确定人类细胞色素P450 2A和2E酶的结构，以扩展和测试我们对这些酶中配体结合的理解，2)开发和应用配体结合人类膜细胞色素P450酶的溶液核磁共振分析，以及3)鉴定和设计有效的，选择性的2A13抑制剂，用于潜在的肺癌化学预防发展。在完成这些研究后，我们期望能够确定2A酶的现有结构是否足以预测新药和小分子的结合，提出抑制剂以减少烟草相关肺癌，并开发和验证溶液核磁共振作为一种有价值的正交技术来研究配体结合。这些结果通过开发询问P450配体结合的新方法间接推进了NIH的目标，并直接导致了减少人类肺癌的新治疗方法。

DESCRIPTION (provided by applicant): Xenobiotic-metabolizing cytochrome P450 (P450) enzymes play key roles in drug metabolism and toxin and procarcinogen activation. Since each P450 can interact with a wide range of substrates and inhibitors, understanding and predicting how foreign compounds bind and are metabolized by these enzymes is of great practical value in toxicology, chemoprevention, and polypharmacy. Building on existing knowledge in the biochemistry and structural biology of human cytochrome P450 2A and 2E1 enzymes, the objective of this proposal is to expand, test, and apply our understanding of the unique relationships between the structures of human 2A and 2E enzymes and their ligand selectivity. It is important to be able to understand the physiological and drug metabolism roles of each enzyme in order to manipulate them to positively affect human health. Relevant to this proposal, 2A13 in the respiratory track activates one of the two most prevalent and carcinogenic compounds in tobacco smoke to produce metabolites that alkylate DNA and can initiate lung cancer. Selective inhibition of 2A13 would significantly reduce this process. Specifically, we propose to 1) determine structures of human cytochrome P450 2A and 2E enzymes with physiologically relevant and marker compounds to both expand and test our understanding of ligand binding in these enzymes, 2) develop and apply a solution NMR assay of ligand binding to human membrane cytochrome P450 enzymes, and 3) identify and design potent, selective inhibitors of 2A13 for potential development as lung cancer chemopreventatives. At the completion of these studies, we expect to be able to determine if existing structures of 2A enzymes are sufficient to predict binding of new drugs and small molecules, to have proposed inhibitors to reduce tobacco- related lung cancer, and to have developed and validated solution NMR as a valuable orthogonal technique to investigate ligand binding. These results advance NIH goals indirectly by developing new methods to interrogate P450 ligand binding and directly by leading to a novel therapeutic approach for the reduction of human lung cancer. PUBLIC HEALTH RELEVANCE: This proposal investigates enzymes involved in the breakdown of drugs and the formation of carcinogens in humans. By understanding how different drugs and foreign chemicals bind and are then broken down by these enzymes, we can better predict how new drugs and known carcinogens are processed and eliminated in the human body. One specific application of this knowledge is the development of a drug that reduces the body's ability to convert nicotine into DNA-damaging molecules that can cause lung cancer in smokers.