Co(II)-Radical Pair Dynamics in B12 Enzyme Catalysis

B12 酶催化中的 Co(II)-自由基对动力学

基本信息

批准号：
7163573
负责人：
KURT WARNCKE
金额：
$ 23.88万
依托单位：
EMORY UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
1998
资助国家：
美国
起止时间：
1998-03-15 至 2007-12-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/7163573
关键词：
Address Amines Apoenzymes Aspirin Binding Biological Biology Carbon Catalysis Cell Nucleus Class Classification Cobalamin Cobalt Coenzymes Complement component C1s Coupled Coupling DNA Sequence Rearrangement DNA biosynthesis Databases Deoxyadenosines Detection Development Electron Spin Resonance Spectroscopy Electrons Enzymes Equilibrium Ethanolamine Ammonia-Lyase Event Family Free Energy Free Radicals Frequencies Generations Genetic Recombination Goals Holoenzymes Hour Hydrogen In Situ Individual Isotopes Kinetics Lasers Lead Ligands Maps Measurement Measures Mediating Metals Methodology Methods Modeling Molecular Nitrogen Nuclear Object Attachment Optics Pharmaceutical Preparations Physiologic pulse Positioning Attribute Principal Investigator Prostaglandin-Endoperoxide Synthase Proteins Pulse taking Range Rate Reaction Research Rest Ribonucleotide Reductase Sampling Site Spectrum Analysis Staging Step Tests Structure System Techniques Testing Therapeutic Thermodynamics Time Work absorption analog base chemical reaction cob(II)alamin cobamamide coenzyme B cofactor comparative corrin covalent bond density deoxyadenosine design enzyme mechanism enzyme substrate analog flash photolysis in vivo insight metalloenzyme migration novel photolysis programs protein structure

项目摘要

Enzymes that harness the extreme reactivity of electron-deficient free radical species carry out some of the most difficult chemical reactions in biology. The regio- and stereo-selectivity achieved by these enzymes defies long-held ideas that radical reactions are non-specific. This class includes the following: ribonucleotide reductases, which catalyze the first unique step in DNA biosynthesis, prostaglandin H- synthase, the target of aspirin and other non-steroidal anti-infiamatory drugs, and the family of coenzyme B12-dependent enzymes, which catalyze metabolite covalent bond rearrangements. The common primary step in the catalyses is metal-assisted generation of an electron-deficient organic radical. This initiator radical, either by itself or through secondary radical species, promotes hydrogen atom abstraction from the substrate to form a substrate-based radical, opening a new reaction channel that facilitates rearrangement to a product radical. An outstanding issue is how the radical pair is stabilized against rapid recombination to achieve productive reaction in high yield. Elucidating the basic principles of how protein and cofactors guide radical stabilization and ensuing substrate radical rearrangement will be sustained focuses of the proposed studies. The adenosylcobalamin-dependent systems, and ethanolamine deaminase specifically, have been selected for scrutiny. The mechanisms of holoenzyme assembly, and radical pair generation, separation and stabilization will be studied by techniques of pulsed-electron paramagnetic resonance and visible/near- infrared absorption spectroscopy by using cryotrapped samples and time-resolved tracking on time scales ranging from picoseconds to hours. The results will be used to construct a detailed molecular mechanism for the enzyme reactions. The insights and novel methods developed will promote identification of radical intermediates in other enzyme reactions, indicate designs for programmed site-specific radical reactions in vivo, and assist therapeutic efforts to combat biologically-destructive free radicals.

利用电子缺陷自由基物种的极端反应性的酶进行了一些生物学中最困难的化学反应。这些酶实现的区域和立体观念违反了根本反应是非特异性的长期以来的想法。该课程包括以下内容：核糖核苷酸还原酶催化了DNA生物合成的第一个独特步骤，前列腺素H- 合酶，阿司匹林和其他非甾体类抗分子药物的靶标，以及辅酶家族 B12依赖性酶，催化代谢产物共价重排。共同的初选在Catalyses中逐步是金属辅助产生电子缺陷的有机自由基。这个发起者自由基本身或通过二级自由基物种，促进了从底物形成基于底物的激进，开设了一个新的反应通道，该通道有助于重排产物激进。一个杰出的问题是如何将自由基对稳定在快速重组中以高产量获得生产反应。阐明蛋白质和辅因子指南的基本原理根治性的稳定和随后的基材自由基重排将是所提出的持续的重点研究。腺苷胆碱素依赖性系统和乙醇胺脱氨酶特定于选择进行审查。全酶组装的机制以及激进的成对产生，分离和稳定将通过脉冲电子顺磁共振和可见/接近 - 的技术研究红外吸收光谱通过使用冷冻样品和时间尺度上的时间分辨跟踪从picseconds到几个小时不等。结果将用于构建详细的分子机制用于酶反应。开发的见解和新方法将促进自由基的识别其他酶反应中的中间体指示了针对编程位点特异性自由基反应的设计体内，并协助治疗努力，以打击生物损害的自由基。