ACTIVE SITES MODEL OF CYTOCHROMES P450 1A1 AND 1A2

描述：（申请人摘要中的逐字记录）人细胞色素P450 1A 1和1A 2在化学物质的代谢活化中起重要作用 致癌物质和药物代谢。特别是P450 1A 1， 存在于肺部，被认为与肺癌有关。这是因为 这种酶氧化高毒性化合物的能力，如多环 芳烃（PAH），其致癌衍生物。P450 1A2，a 典型的肝脏P450，在药物代谢中起作用，并催化 杂环胺代谢活化为遗传毒性产物。为了 设计有效的化疗药物， 癌症，重要的是要阐明这些结构-功能关系 内切酶 细胞色素P450 1A 1和1A 2在序列上非常相似（72 同一性），但底物和抑制剂特异性不同。一般来说，1A 1 表现出对多环芳烃，例如苯并（a）芘， 而1A 2与杂环胺底物如咖啡因有关。 拟议研究的长期目标是阐明 这些差异的基础。要检验的假设是， 这些酶的结构是相似的，功能差异是由 控制酶的关键氨基酸残基的离散数量的存在 功能在一种酶中取代这些残基以模拟另一种酶， 相互转化的活动。这将通过酶的分子建模来测试 结构与实验方法相结合，如现场指导 诱变和异源表达。三维酶模型将 用于鉴定负责独特底物的关键氨基酸残基 P450 1A 1和1A 2的特异性和抑制剂敏感性。分析 酶-底物相互作用和分子动力学模拟将有助于 这两种酶之间功能差异的解释。建模 预测将遵循的建设，表达和功能 评估适当的突变蛋白，实验结果将是 用于在迭代方法中细化模型。的创新特点 建议包括广泛利用分子动力学模拟， 结合自由能计算以预测底物特异性和抑制剂 选择性该项目的多学科性质，它结合了两个 显著不同的研究领域，如计算化学和 生物化学/分子生物学，应该证明在研究 复杂的生化和毒理学问题。拟议的研究应 对P450 1A结构及其与 酶的功能，从而为合理设计抗癌药物提供依据 毒品

DESCRIPTION: (Verbatim from the Applicant's Abstract) Human cytochromes P450 1A1 and 1A2 play an important role in the metabolic activation of chemical carcinogens and in the metabolism of drugs. In particular, P450 1A1, which is present in lungs, is thought to be linked to lung cancer. This is because of the enzyme's ability to oxidize highly toxic compounds, such as polycyclic aromatic hydrocarbons (PAHs), to their carcinogenic derivatives. P450 1A2, a typical hepatic P450, plays a role in the metabolism of drugs and catalyzes metabolic activation of heterocyclic amines to genotoxic products. In order to design effective chemotherapeutic agents that would decrease the incidence of cancer, it is important to elucidate structure-function relationships of these enzymes. Cytochromes P450 1A1 and 1A2 are very similar in sequence (72 percent identity), but differ in substrate and inhibitor specificities. In general, 1A1 exhibits a preference for polyaromatic hydrocarbons, such as benzo(a)pyrene, whereas 1A2 is associated with heterocyclic amine substrates, such as caffeine. The long-term objective of the proposed research is to elucidate the structural basis for those differences. The hypothesis to be tested is that the overall structure of these enzymes is similar and functional differences are caused by the presence of a discrete number of key amino acid residues that govern enzyme function. Substitution of these residues in one enzyme to mimic the other will interconvert activities. This will be tested by molecular modeling of enzyme structure coupled with experimental approaches, such as site-directed mutagenesis and heterologous expression. The 3-dimensional enzyme models will be used to identify key amino acid residues responsible for unique substrate specificities and inhibitor susceptibilities of P450 1A1 and 1A2. The analysis of enzyme-substrate interactions and molecular dynamics simulations will aid in the explanation of functional differences between the two enzymes. Modeling predictions will be followed by the construction, expression and functional evaluation of appropriate mutant proteins, and experimental results will be used to refine models in an iterative approach. Innovative features of the proposal include extensive utilization of molecular dynamics simulations and binding free energy calculations to predict substrate specificity and inhibitor selectivity. The multidisciplinary nature of the project, which combines two markedly different areas of investigation, such as computational chemistry and biochemistry/molecular biology, should prove highly advantageous in studies of complex biochemical and toxicological issues. The proposed research should yield valuable insight into the P450 1A structure and its relationship to enzyme function, and thus provide a basis for the rational design of anticancer drugs.