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.

在该项目中得到了无机，生物有机和有机金属化学计划的支持，犹他州立大学的理查德·C·霍尔兹（Richard C. Holz）博士将研究对多核水解酶在催化反应中发挥的单个金属离子在其催化反应中发挥的作用。 该作品的初始阶段将围绕来自Vibrio proteytica的双核锌（II）氨基肽酶的研究建立，为此，可以使用质粒来过表达大肠杆菌中的酶。 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，这些锌离子也已被证明也显示出增强的酶活性。 在某些研究中，镉（II）也将用于取代。 要使用的方法包括UV-VIS，NMR，EPR和EXAFS光谱镜以及稳态动力学。 要考虑的特定问题包括：1）用其他金属离子取代时配体的性质和几何形状是什么，以便获得多动症。 2）每个金属中心在催化中扮演什么角色？ 3）底物如何与双核活性位点相互作用？ 4）一种或两个金属离子是否用作刘易斯酸来结合和激活水分子的水解反应？ 含有多种金属离子（多核酶）的水解酶才证明是最近才存在的。 这些酶作用催化的机制目前尚不清楚。 这项研究旨在确定单个金属离子在催化反应中的作用。 结果可能导致可用于生物技术的合成酶的发展。