Structural Basis of Cytochrome P450 Activity

描述（由申请方提供）：异源代谢细胞色素P450（P450）酶在药物代谢、毒素和原致癌物活化中起关键作用。由于每个P450可以与各种底物和抑制剂相互作用，因此了解和预测外源化合物如何结合并被这些酶代谢，在毒理学，化学预防和多药治疗中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，本提案的目的是扩展，测试和应用我们对人类2A和2E酶结构及其配体选择性之间独特关系的理解。重要的是要能够了解每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。与该提议相关的是，呼吸道中的2A13激活烟草烟雾中两种最普遍和致癌的化合物之一，产生使DNA烷基化的代谢物，并可引发肺癌。选择性抑制2A13将显著减少该过程。具体而言，我们建议1）用生理学相关和标记化合物确定人细胞色素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）开发和应用配体与人膜细胞色素P450酶结合的溶液NMR测定，以及3）鉴定和设计有效的，2A13的选择性抑制剂作为肺癌化学预防剂的潜在发展。在完成这些研究时，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的肺癌，并开发和验证溶液NMR作为研究配体结合的有价值的正交技术。这些结果通过开发新的方法来间接地询问P450配体结合，并直接通过导致一种新的治疗方法来减少人类肺癌，从而推动了NIH的目标。

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.