CAREER: Metalloenzyme mechanisms probed by resonance Raman spectroscopy

金属是约50%蛋白质的重要组成部分，具有从能量储存和代谢到分子运输和细胞信号传导的功能。然而，由于这些金属蛋白系统的复杂性，其中许多包含多个不同的金属中心，其作用机制往往是难以捉摸的。本研究将通过实验室实验与计算化学相结合，开发一种新的方法来研究这些反应，以更好地了解这些金属蛋白的功能。该方法将应用于天然和工程酶的产氢和氧化研究，未来可能应用于替代能源、可持续燃料、二氧化碳固定和氮循环等领域。此外，本提案的研究目标与教育和推广活动相结合，对科学和社会都具有深远的影响。这项工作的跨学科性质，涉及生物学、化学和物理学的各个方面，使这一主题能够为各种各样的受众所接受。该教育计划将以一种身临其境的方式，让来自未被充分代表的种族/民族和残疾的广泛学生接触现代化学研究，目的是建立一个多样化的、以职业为导向的STEM人才管道。将为成人观众举办一般兴趣研讨会，目的是提高科学素养，提高认识，并更好地使社区参与俄勒冈州立大学和世界各地正在进行的当代研究。来自俄亥俄州立大学的Hannah Shafaat博士将利用计算引导共振拉曼光谱（CGRRS）研究金属蛋白中的酶促机制，并获得化学学部生命过程化学项目的奖项。通过利用共振拉曼光谱的选择性和灵敏度，在一个高度可调的、最先进的仪器上进行，将获得催化中间体结构的重要信息，包括质子化状态、键序和配位模式。位点选择性和状态特异性激发结合光谱特性的理论预测将用于表征中间体和瞬态物质，这是传统方法无法达到的。将研究金属酶活性位点的小分子活化反应，包括氢化酶和模型中的质子还原、氢氧化和氧还原，以证明这种实验计算方法的实用性，并确定催化过程中的关键结构基元。这种协同方法将广泛适用于多种酶的研究，特别是那些含有多种金属辅助因子的酶，如氮酶和一氧化碳脱氢酶。使用CGRRS，将有可能获得分子水平上对复杂反应机制的洞察，并开始了解金属蛋白如何能够作为一些已知的最有效的催化剂发挥作用。

Metals are an essential component of approximately 50% of all proteins, with functions ranging from energy storage and metabolism to molecular transport and cellular signaling. However, due to the complexity of these metalloprotein systems, many of which contain multiple, distinct metal centers, the mechanisms of action often remain elusive. This research will develop a novel approach towards studying these reactions by combining laboratory experiments with computational chemistry to better understand how these metalloproteins function. This methodology will be applied to the study of hydrogen production and oxidation in natural and engineered enzymes, with future potential applications for alternative energy, sustainable fuels, carbon dioxide fixation, and nitrogen cycling, among others. Furthermore, the research objectives of this proposal are integrated with education and outreach activities, with far-reaching impact for both science and society. The interdisciplinary nature of the work, involving aspects of biology, chemistry, and physics, renders the subject accessible to a wide variety of audiences. The educational plan will expose a broad range of students from underrepresented racial/ethnic groups and with disabilities to modern chemistry research in an immersive fashion, with the objective of building a diverse, career-oriented pipeline of STEM talent. General-interest seminars for adult audiences will be presented with aims of improving scientific literacy, increasing awareness, and better engaging the community in contemporary research being carried out at OSU and across the world.With this award from the Chemistry of Life Processes Program in the Division of Chemistry, Dr. Hannah Shafaat from The Ohio State University will study enzymatic mechanisms in metalloproteins using computationally-guided resonance Raman spectroscopy (CGRRS). By harnessing the selectivity and sensitivity of resonance Raman spectroscopy performed on a highly tunable, state-of-the-art instrument, important information on the structures of catalytic intermediates, including protonation state, bond order, and coordination mode, will be obtained. Site-selective and state-specific excitation in conjunction with theoretical predictions of spectroscopic properties will be used to characterize intermediates and transient species that are inaccessible by conventional approaches. Small molecule activation reactions at metalloenzyme active sites, including proton reduction, hydrogen oxidation, and oxygen reduction in hydrogenases and models, will be investigated to demonstrate the utility of this experimental-computational approach and identify key structural motifs during catalysis. This synergistic methodology will be broadly applicable to the study of many classes of enzymes, particularly those containing multiple metallocofactors, such as nitrogenase and carbon monoxide dehydrogenase. Using CGRRS, it will be possible to gain molecular-level insight into complex reaction mechanisms and begin to understand how metalloproteins are able to function as some of the most efficient catalysts known.