Enzyme Structure-Function Relationships

酶结构与功能的关系

• 批准号: 9118302
• 负责人: Stephen Remington
• 金额: $ 29.4万
• 依托单位: University of Oregon Eugene
• 依托单位国家: 美国
• 项目类别: Continuing Grant
• 财政年份: 1992
• 资助国家: 美国
• 起止时间: 1992-03-01 至 1995-08-31
• 项目状态: 已结题
• 来源: https://www.nsf.gov/awardsearch/showAward?AWD_ID=9118302&HistoricalAwards=false
• 关键词: Enzyme; Structure; Function; Relationships

Two approaches are proposed to investigate substrate specificity and catalytic mechanisms of two different enzymes, citrate synthase (CS) and serine carboxypeptidase (CPD). The first will be an X-ray crystallographic analysis of the structures of the enzymes and to compare ligand affinities and catalytic activity of related or mutant forms of the enzymes. The second will be to investigate the role of electrostatic fields in the mechanisms by use of theoretical calculations. Citrate synthase is one of few enzymes capable of directly forming a carbon bond. A detailed mechanism for part of the reaction has been proposed. Site-directed mutants of CS with differing activity have been constructed. Their structures will be analyzed by crystallographic techniques, enzyme kinetics and electrostatic field calculations to study the roles of selected side chains. Serine carboxypeptidases are highly specific proteinases which are essential for the maturation of peptide hormones. They are unrelated to other serine proteinases, but have the same active site groups. Analysis of inhibitor binding will determine the basis for substrate specificity. Selected mutants of yeast CPD will be studied to determine the basis for the unusual pH dependence of the reaction and to test whether the enzymatic mechanism is similar to that proposed for other serine proteinases.

提出了两种研究底物特异性的方法 柠檬酸合成酶和柠檬酸合成酶两种不同的酶的催化机制 (CS)和丝氨酸羧肽酶（CPD）。 第一个是X光 酶结构的晶体学分析， 比较配体亲和力和催化活性的相关或 酶的突变形式。 第二个将是调查 静电场的作用机制，通过使用 理论计算。 柠檬酸合成酶是少数几种能够直接形成 碳键 该反应的一部分的详细机理已经 提出如 具有不同活性的CS定点突变体 已经建成。 它们的结构将通过 晶体学技术、酶动力学和静电 现场计算，以研究选定的侧链的作用。 丝氨酸羧肽酶是高度特异性的蛋白酶， 对肽激素的成熟至关重要。 他们是 与其他丝氨酸蛋白酶无关，但具有相同的活性 站点组。 抑制剂结合的分析将确定 底物特异性的基础。 酵母CPD的选定突变体 将进行研究，以确定不寻常的pH值的基础 反应的依赖性，并测试酶是否 其机制类似于对其他丝氨酸蛋白酶提出的机制。