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

描述（由申请人提供）： 利用缺电子自由基物种的极端反应性的酶进行生物学中一些最困难的化学反应。这些酶所实现的区域和立体选择性挑战了长期以来认为自由基反应是非特异性的想法。本课程包括以下内容：核糖核苷酸还原酶，催化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抗炎药的靶标，S-腺苷甲硫氨酸依赖性酶，具有超过六百个成员，催化广泛的自由基介导的氧化还原反应，以及辅酶B12（腺苷钴胺素）依赖性酶超家族，其成员催化代谢物共价键重排。在这些化学性质不同的催化剂中，共同的主要步骤是金属辅助产生缺电子有机自由基。该引发剂自由基本身或通过第二自由基物质促进氢原子从底物中提取以形成基于底物的自由基，从而打开促进转化为产物的新反应通道。一个突出的问题是如何使自由基对稳定以防止快速重组，从而以高产率实现生产性反应。阐明蛋白质和辅因子如何引导自由基产生、稳定、蛋白质内自由基迁移和自由基重排的基本原理将是拟议研究的持续重点。辅酶B12依赖性酶，特别是乙醇胺氨裂解酶，已被选定进行审查。分子结构和动力学对酶功能的贡献将通过使用脉冲电子顺磁共振和可见光/近红外吸收光谱技术，在冷冻和时间分辨系统的时间尺度从皮秒到小时进行研究。开发的基本见解和新方法将促进其他生物反应中自由基中间体的识别和表征，为程序化自由基反应性的设计提供信息，并协助分子治疗工作来对抗有害的自由基过程。公共卫生相关性：利用缺电子自由基的极端反应性的酶可以执行生物学中一些最困难的反应。我们试图了解分子结构和动力学的贡献，这种生产性的自由基反应辅酶B12依赖性酶通过使用光谱技术。开发的基本见解和新方法将促进其他生物系统中自由基中间体的表征，告知程序化自由基反应性的设计，并协助分子治疗努力对抗有害的自由基过程。

DESCRIPTION (provided by applicant): 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-inflammatory drugs, the S-adenosylmethionine-dependent enzymes, with over six-hundred members that catalyze a wide range of radical- mediated redox reactions, and the coenzyme B12 (adenosylcobalamin)-dependent enzyme superfamily, whose members catalyze metabolite covalent bond rearrangements. The common primary step in these chemically-disparate 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 transformation to the product. 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 cofactor guide radical generation, stabilization, intra-protein radical migration, and radical rearrangement will be sustained focuses of the proposed studies. The coenzyme B12-dependent enzymes, and ethanolamine ammonia-lyase specifically, have been selected for scrutiny. The contributions of molecular structure and dynamics to enzyme function will be studied by using techniques of pulsed-electron paramagnetic resonance and visible/near-infrared absorption spectroscopy, in cryotrapped and time-resolved systems on time scales ranging from picoseconds to hours. The fundamental insights and novel methods developed will promote identification and characterization of radical intermediates in other biological reactions, inform the design of programmed radical reactivity, and assist molecular-therapeutic efforts to combat pernicious free radical processes. PUBLIC HEALTH RELEVANCE: Enzymes that harness the extreme reactivity of electron-deficient free radicals perform some of the most difficult reactions in biology. We seek to understand the contributions of molecular structure and dynamics to this productive radical reactivity in coenzyme B12-dependent enzymes by using spectroscopic techniques. The fundamental insights and novel approaches developed will promote characterization of radical intermediates in other biological systems, inform the design of programmed radical reactivity, and assist molecular- therapeutic efforts to combat pernicious free radical processes.