SPECTROSCOPIC STUDIES OF ENZYME SUBSTRATE COMPLEXES

酶底物复合物的光谱研究

• 批准号: 6309211
• 负责人: GEORGE H REED
• 金额: $ 0.75万
• 依托单位: UNIVERSITY OF WISCONSIN-MADISON
• 依托单位国家: 美国
• 财政年份: 2000
• 资助国家: 美国
• 起止时间: 2000-04-15 至 2005-02-28
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6309211
• 关键词: biological products; biomaterials; biomedical resource; enzymes; nuclear magnetic resonance spectroscopy

The structures of complexes of yeast enolase and rabbit muscle pyruvate kinase have been solved to high resolution by X-ray crystallography. These structures reveal the identities of active-site residues in these two enzymes. Site-directed mutagenesis has been performed to clarify the roles of the active-site residues in catalysis. Residues believed to function as the acid/base catalysts for these enzymes are lysine 345 for yeast enolase and either arginine 72 or threonine 327 for pyruvate kinase. Both enzymes catalyze alternate reactions. NMR characterization of the products of these alternate reactions will be carried to help corroborate our hypotheses. NMR identification of the stereospecificity of reactions catalyzed by these enzymes and mutants will also be employed.

酵母烯醇化酶与兔肌肉复合物的结构 丙酮酸激酶已通过 X 射线高分辨率解析 晶体学。 这些结构揭示了 这两种酶的活性位点残基。 定点诱变 已进行以澄清活性位点残基的作用 催化。 据信具有酸/碱催化剂作用的残留物 对于这些酶，酵母烯醇化酶的赖氨酸 345 和精氨酸 72 或苏氨酸 327 用于丙酮酸激酶。 两种酶都催化 交替反应。 这些产品的 NMR 表征 将进行替代反应以帮助证实我们的 假设。 核磁共振鉴定反应的立体专一性 也将使用由这些酶和突变体催化的酶。