Defining the mechanistic determinants of catalytic bias in cofactor-based enzymatic oxidation-reduction reactions

定义基于辅因子的酶促氧化还原反应中催化偏差的机械决定因素

• 批准号: 10874184
• 负责人: JOHN W PETERS
• 金额: $ 31.26万
• 依托单位: UNIVERSITY OF OKLAHOMA
• 依托单位国家: 美国
• 财政年份: 2020
• 资助国家: 美国
• 起止时间: 2020-09-15 至 2024-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10874184
• 关键词: Defining; mechanistic; determinants; catalytic; bias

PROJECT SUMMARY Enzymatic reactions comprise the basic building blocks essential to the biochemical processes of cellular metabolism. While most enzymatic reactions are reversible, certain enzymes accelerate a reaction in one direction significantly differently than the reverse direction. The mechanistic determinants of this phenomenon – often referred to as “catalytic bias” – are generally unknown. Our long-term goal of this project is to identify the fundamental principles adopted by redox enzymes (enzymes involved in oxidation-reduction reactions) that provide molecular level control of the catalytic bias. In doing so, we will provide a greater fundamental understanding of the factors that control metabolic processes in all life. Specifically, the objective of this proposal is to delineate determinants that influence the directional reactivity of model Fe containing hydrogenases that are structurally related but exhibit large differences in catalytic bias for the specific reversible hydrogen (H2) oxidation reaction from electron and protons. The central hypothesis is that catalytic bias commonly in redox enzymes results from the relative differential stabilization and destabilization of oxidation states of the respective redox cofactors critical to determining the rate limiting step of the catalytic cycle. The rationale underlying the proposal is that the implementation will result in defining mechanistic determinants of the proposed model system. The proposed work will culminate in elucidating universal, mechanistic factors of catalytic bias that govern a large number of, if not all, redox enzymes. The central hypothesis will be tested by pursuing three specific aims that examine how 1) the reduction potential of the active site proximal accessory redox clusters; 2) the characteristics of the active site cluster protein environment; and 3) the secondary coordination sphere structural dynamics can control reactivity and catalytic bias in cofactor-based oxidation reduction catalysis. To pursue these aims, we will employ an innovative strategy integrating biochemical, structural, spectroscopic, and computational approaches on a unique, one-of-a-kind model hydrogenase platform. The proposed research is significant, because it will delineate molecular determinants that influence the fine-tuning of redox cofactors through the relative stabilization or destabilization of oxidation states to afford certain reactivity fundamental to directing energy and matter in cells. The work will provide additional foundational resources in the form of a blueprint for integrating multiple biophysical and computational approaches.

项目概要 酶促反应包含细胞生化过程所必需的基本构件 代谢。虽然大多数酶促反应是可逆的，但某些酶会加速一种反应 方向与相反方向明显不同。这种现象的机械决定因素—— 通常被称为“催化偏差”——通常是未知的。我们这个项目的长期目标是确定 氧化还原酶（参与氧化还原反应的酶）采用的基本原理 提供催化偏向的分子水平控制。在此过程中，我们将为 了解控制所有生命代谢过程的因素。具体而言，本提案的目标 是为了描绘影响含铁模型氢化酶定向反应性的决定因素 结构相关，但对特定可逆氢 (H2) 的催化偏差表现出巨大差异 电子和质子的氧化反应。中心假设是氧化还原反应中普遍存在催化偏向 酶是由各自氧化态的相对差异稳定和不稳定造成的 氧化还原辅因子对于确定催化循环的限速步骤至关重要。其背后的基本原理 提议是，实施将导致定义提议模型的机械决定因素 系统。拟议的工作最终将阐明催化偏差的普遍性、机械性因素，这些因素 控制大量（如果不是全部）氧化还原酶。中心假设将通过追求三个 具体目标是检查如何 1) 活性位点近端辅助氧化还原簇的还原潜力； 2） 活性位点簇蛋白环境的特征； 3）次要协调圈 结构动力学可以控制基于辅因子的氧化还原催化中的反应性和催化偏差。到 为了实现这些目标，我们将采用整合生化、结构、光谱和 独特的、独一无二的模型氢化酶平台上的计算方法。拟议的研究是 意义重大，因为它将描绘影响氧化还原辅助因子微调的分子决定因素 通过氧化态的相对稳定或不稳定来提供某些基本的反应性 引导细胞内的能量和物质。这项工作将以以下形式提供额外的基础资源： 整合多种生物物理和计算方法的蓝图。