Co(II) Radical Pair Dynamics in B12 Enzyme Catalysis

描述（由申请人提供）：拟议研究的长期目标是阐明在金属自由基对生成和耦合自由基介导反应的背景下，控制酶催化的基本分子原理。这一目标将通过对鼠伤寒沙门氏菌的辅酶B12（腺苷钴胺素）依赖性酶，乙醇胺氨裂解酶的生物化学和物理研究的综合计划来实现。我们已经开发了新的方法，软件和硬件的脉冲电子顺磁共振（EPR）光谱。结合生物化学技术，我们开发了低温系统和时间分辨EPR方法，这些方法能够实现开拓性的实验努力，以单步分辨率直接解决乙醇胺氨裂解酶中的共价键断裂/键形成反应。总体目标是定义引导反应的自由能景观，确定起源并表征蛋白质和耦合溶剂波动的作用，这些波动推动系统在这一景观上，并解析构建自由能景观的埃尺度分子结构。该项目提出了一个酶催化的“构型催化”模型，并寻求建立自由基重排作为定义酶反应中化学步骤的景观和蛋白质-溶剂运动贡献的范例。溶剂的结构和动力学的酶反应的贡献也解决了一套EPR光谱和弛豫技术。研究结果将影响生物医学和人类健康的分子方法，包括抗微生物疗法，催化剂，抑制剂和生物纳米室的合理设计，以及自由基反应的控制。

DESCRIPTION (provided by applicant): The long-term goal of the proposed research is to elucidate fundamental molecular principles that govern catalysis in enzymes, in the context of metal-radical pair generation and coupled radical-mediated reactions. This goal will be enacted through a comprehensive program of biochemical and physical studies of the coenzyme B12 (adenosylcobalamin) - dependent enzyme, ethanolamine ammonia-lyase, from Salmonella typhimurium. We have developed new methods, software and hardware for pulsed-electron paramagnetic resonance (EPR) spectroscopy. In conjunction with biochemical techniques, we have developed low-temperature systems and time- resolved EPR approaches that enable pioneering experimental efforts that address, directly, covalent bond- breaking/bond-making reactions in ethanolamine ammonia-lyase at single-step resolution. The general aims are to define the free energy landscape that guides the reactions, identify the origins and characterize the roles of the protein and coupled solvent fluctuations that propel the system over this landscape, and resolve the Angstrom-scale molecular structures that frame the free energy landscape. The projects develop a proposed "configurational catalysis" model for enzyme catalysis, and seek to establish the radical rearrangement as a paradigm for defining landscape and protein-solvent motional contributions to chemical steps in enzyme reactions. Contributions of solvent structure and dynamics to the enzyme reactions are also addressed by a suite of EPR spectroscopic and relaxation techniques. The outcomes of the research will impact molecular approaches in biomedicine and human health, including anti- microbial therapies, rational design of catalysts, inhibitors, and bio-nanocompartments, and control of free radical reactions.