CHEMICAL MECHANISMS OF BIOSYNTHESIS

生物合成的化学机制

• 批准号: 6385012
• 负责人: RICHARD VANCE WOLFENDEN
• 金额: $ 27.34万
• 依托单位: UNIV OF NORTH CAROLINA CHAPEL HILL
• 依托单位国家: 美国
• 财政年份: 1980
• 资助国家: 美国
• 起止时间: 1980-07-01 至 2004-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6385012
• 关键词: active sites; aminohydrolases; binding sites; biosynthesis; decarboxylases; enzyme activity; enzyme inhibitors; enzyme mechanism; enzyme substrate analog; enzyme substrate complex; protein binding

DESCRIPTION: (adapted from applicant's abstract) As a substrate progresses from the ES complex to the transition state, binding grows stronger by a factor that matches or surpasses the rate enhancement. The principal investigator recently began to explore the rate enhancements that enzymes produce, by measuring the rates of the corresponding reactions in the absence of a catalyst. The results offer a means of recognizing those enzymes that produce the largest rate enhancements, a basis for analyzing the effects of enzyme modification, and a guide to those enzymes that should serve as the most sensitive targets for inhibitor design, useful in designing new transition state analogues as drugs. Dr. Wolfenden proposes to extend those measurements to include each of the major classes of reactions catalyzed by enzymes. In addition to very strong binding of the altered substrate in the transition state, a major challenge faced by an enzyme's active site is to avoid tight binding of the substrate in the ground state. To understand this aspect of enzyme action, he is also investigating the structures of enzyme-product complexes, for comparison with those of enzyme complexes with S* analogues. Finally, he seeks to determine the nature of the detailed forces that are involved in S* binding, which constitute an enzyme's mechanism of action. To determine the nature of those forces, he will continue the structural analysis of ES* complexes, focusing on native and mutant E. coli cytidine deaminase and yeast OMP decarboxylase, both expressed in E. coli. The contributions of active site binding determinants to transition state affinity will be evaluated by examining at different temperatures, the kinetic consequences of truncating the enzyme, substrate or transition state analogue, to evaluate individual group contributions to the free energy, enthalpy and entropy of enzyme-substrate association in the ground state and transition state. In addition, comparison of the transition state binding properties of the native enzyme with those its pieces, obtained by mutation, is expected to reveal the benefit to catalysis that an enzyme gains from being properly connected.

描述：（根据申请人的摘要改编）基板从 ES复合物到过渡状态，结合增强了一个因素 匹配或超过速率提高。主要调查员 通过测量 在没有催化剂的情况下，相应反应的速率。结果 提供一种识别那些产生最大速率的酶 增强功能，分析酶修饰作用的基础和A 那些应该用作最敏感靶标的酶的指南 抑制剂设计，可用于设计新的过渡状态类似物作为药物。 沃尔芬登博士建议将这些测量扩展到包括每个测量 酶催化的主要反应类别。除了很强 在过渡状态下改变的底物的结合，这是一个重大挑战 酶的活性位点面对的是避免底物紧密结合 基态。要了解酶作用的这一方面，他也是 研究酶产物复合物的结构，以与 具有S*类似物的酶复合物。最后，他试图确定 构成S*结合所涉及的详细力的性质 酶的作用机理。为了确定这些力量的性质，他 将继续对ES*复合物进行结构分析，重点是本地和 突变的大肠杆菌胞苷脱氨酶和酵母OMP脱羧酶都表达 在大肠杆菌中。主动位点结合因素对过渡的贡献 国家亲和力将通过在不同温度下检查， 截断酶，底物或过渡状态的动力学后果 模拟，评估各个群体对自由能的贡献， 焓和基底酶的熵在基态和 过渡状态。另外，过渡状态结合的比较 通过突变获得的天然酶的特性，其碎片是 有望揭示催化的好处，即酶从 正确连接。