Mechanistic Studies on Dinuclear Hydrolytic Catalysts

双核水解催化剂的机理研究

• 批准号: 9422098
• 负责人: Richard Holz
• 金额: $ 24.03万
• 依托单位: Utah State University
• 依托单位国家: 美国
• 项目类别: Continuing Grant
• 财政年份: 1995
• 资助国家: 美国
• 起止时间: 1995-04-01 至 1998-03-31
• 项目状态: 已结题
• 来源: https://www.nsf.gov/awardsearch/showAward?AWD_ID=9422098&HistoricalAwards=false
• 关键词: Mechanistic; Studies; Dinuclear; Hydrolytic; Catalysts

In this project, which is supported by the Inorganic, Bioinorganic and Organometallic Chemistry Program, Dr. Richard C. Holz of Utah State University will undertake a study of the roles that the individual metal ions in multinuclear hydrolytic enzymes play in their catalytic reactions. The initial phase of the work will be built around studies of the dinuclear Zn(II) aminopeptidase from Vibrio proteolytica, for which a plasmid is available that can be used to overexpress the enzyme in E. coli. The approach will involve substitution of individual zinc ions in the enzyme with spectroscopically active Co(II), Ni(II) or Cu(II) ions, which have already been demonstrated to also show enhanced enzymatic activity. Cadmium(II) will also be used for substitution in some studies. Methods to be used include UV-VIS, NMR, EPR and EXAFS spectroscopies, as well as steady state kinetics. Specific questions to be considered include: 1) What is the nature and geometry of the ligands upon substitution with other metal ions such that hyperactivity is obtained. 2) What specific role does each metal center play in catalysis? 3) How do substrates interact with the dinuclear active site? and 4) Does one or both metal ions serve as a Lewis acid to bind and activate a water molecule for the hydrolysis reaction? Hydrolytic enzymes that contain more than one metal ion (multinuclear enzymes) have been demonstrated to exist only recently. The mechanisms by which these enzymes effect catalysis are presently unknown. This research is designed to determine the role that the individual metal ions play in the catalytic reaction. The results may lead to the development of synthetic enzymes that can be used in biotechnology.

在这个项目中，这是由无机，生物无机和有机化学计划，博士理查德C。犹他州州立大学的霍尔茨将进行一项研究，研究多核水解酶中单个金属离子在催化反应中所起的作用。 这项工作的初始阶段将围绕来自溶蛋白弧菌的双核Zn（II）氨肽酶的研究进行，对于这种酶，可以使用质粒在E.杆菌 该方法将涉及用光谱活性Co（II）、Ni（II）或Cu（II）离子取代酶中的单个锌离子，这些离子已经被证明也显示出增强的酶活性。 在某些研究中，镉（II）也将被用于替代。 使用的方法包括UV-VIS、NMR、EPR和EXAFS光谱，以及稳态动力学。 需要考虑的具体问题包括：1）当被其他金属离子取代从而获得超活性时，配体的性质和几何形状是什么。 2)每个金属中心在催化作用中起什么特定的作用？ 3)底物如何与双核活性中心相互作用？ 和4）一种或两种金属离子是否用作刘易斯酸以结合和活化水分子用于水解反应？ 含有一个以上金属离子的水解酶（多核酶）最近才被证明存在。 这些酶影响催化作用的机制目前尚不清楚。 本研究旨在确定单个金属离子在催化反应中的作用。 这些结果可能会导致可用于生物技术的合成酶的开发。