Complementary Studies of Native Sulfite Reductase and Biosynthetic Model to Understand Structural Features Responsible for Selective Multi-Electron Reduction of Sulfite

天然亚硫酸盐还原酶和生物合成模型的互补研究，以了解负责亚硫酸盐选择性多电子还原的结构特征

• 批准号: 10330437
• 负责人: Christopher (Chris) J Reed
• 金额: $ 1.43万
• 依托单位: UNIVERSITY OF TEXAS AT AUSTIN
• 依托单位国家: 美国
• 财政年份: 2020
• 资助国家: 美国
• 起止时间: 2020-02-16 至 2022-04-10
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10330437
• 关键词: Aconitate Hydratase; Active Sites; Address; Animals; Assimilations; Binding; Binding Sites; Biology; Catalysis; Chemicals; Chemistry; Complex; Coupled; Drug Metabolic Detoxication; Electrons; Enzymes; Ferredoxin; Goals; Health; Healthcare; Heme; Hemeproteins; Human; Knowledge; Lead; Learning; Ligands; Metals; Modeling; Modification; Molecular; Nature; Oxidation-Reduction; Pathogenicity; Property; Proteins; Protocols documentation; Protons; Reaction; Reagent; Reporting; Research; Research Training; Respiration; Role; Scaffolding Protein; Site; Structure; Structure-Activity Relationship; Sulfides; Sulfite reductase; Sulfites; Sulfur; System; Techniques; Testing; Transition Elements; Variant; Virulence; Work; X-Ray Crystallography; base; career; cofactor; design; enzyme substrate; experimental study; heme C; improved; metalloenzyme; microbial; microorganism; novel; pathogen; scaffold; training opportunity

Project Summary/Abstract The goal of this project is to obtain detailed understanding of the structural features present in a class of multinuclear metalloenzymes that make it capable of promoting selective 6 e- reduction of sulfite (SiRs), and address important scientific issues in the fields of complex multinuclear transition metal active sites and the global sulfur cycle. Specifically, this project seeks to understand the role of the coupled auxiliary [4Fe-4S] cluster reduction potential on facilitating efficient—and selective—S-O bond cleavage of sulfite, along with the chemical nature of intermediates that have been proposed in the catalytic cycle of SiRs. To achieve this goal, this project proposes complimentary studies, both of a native SiR protein, and a biosynthetic model capable of catalyzing sulfite reduction—the only current structural and functional model of SiRs. Using a novel biosynthetic approach, which aims to overcome limitations inherent to studies of the native enzyme alone, a robust scaffold will be used to learn which features of SiR are crucial for its activity, through experiments designed to display a gain in function, as opposed to inferring structure-function relationship through loss of activity in the native enzyme. The proposed complimentary studies of these two systems will allow us to identify the structural features unique to SiRs that lead to high catalytic efficiency towards a complex multi-electron, multi-proton transformation. We will be able to understand: (1) the importance of the redox active [4Fe-4S] cluster coupled to the siroheme substrate binding site on SiR activity, (2) the nature of postulated Fe-SOx intermediates, and their relevance to the catalytic cycle of SiR, and (3) the features of the siroheme ligand in SiR which make it suitable for sulfite reduction activity, relative to the heme c of the biosynthetic scaffold. Achieving the above goals will result in a deeper understanding of the structure and function of SiRs that may be difficult to achieve by studying the native enzyme alone. By mimicking the function of a native system through rational modifications with a different protein scaffold, generalizable conclusions can be made concerning important structural features of the native active site. This research plan will advance the knowledge of a broad range of multinuclear metalloenzymes relevant to human health, specifically, related to their structure, function, and metalloenzyme design in general, while providing an excellent training opportunity to achieve the career goal the candidate.

项目摘要/摘要 该项目的目的是获得对目前的结构特征的详细了解 在一类多核金属酶中，能够促进选择性6 e- 减少亚硫酸盐（SIRS），并解决复杂领域的重要科学问题 多核过渡金属活性位点和全局硫周期。具体来说，这个项目寻求 了解耦合辅助[4FE-4S]群集降低潜力对促进的作用 硫酸盐的高效和选择性 - S-O键裂解以及化学性质 在SIR的催化循环中提出的中间体。为了实现这一目标，这个 项目提案免费研究，包括本机SIR蛋白和生物合成模型 能够催化亚硫酸盐还原 - SIR的唯一当前结构和功能模型。 使用一种新型的生物合成方法，旨在克服对研究的局限性 仅本地酶，将使用强大的脚手架来学习爵士的哪些特征至关重要 为了其活动，通过旨在显示功能增长的实验，而不是推断 结构 - 功能通过在天然酶中的活性丧失。提议 对这两个系统的免费研究将使我们能够确定独特的结构特征 向导致高催化效率的SIRS朝着复杂的多电子多物种 转型。我们将能够理解：（1）氧化还原活动的重要性[4FE-4S] 在SIR活动上与Siroheme底物结合位点耦合的簇，（2）假定的性质 Fe-Sox中间体及其与SIR催化周期的相关性，（3） Sir中的siroheme配体使其适用于硫酸盐还原活性，相对于血红素C 生物合成支架。 实现上述目标将使对结构和功能有更深入的了解 单独研究天然酶可能难以实现的SIR。通过模仿 通过具有不同蛋白质支架的理性修饰，天然系统的功能， 可以得出有关本地重要结构特征的可概括结论 活性站点。该研究计划将促进广泛的多核知识 与人类健康相关的金属酶，特别与其结构，功能和 金属酶设计一般，同时提供了极好的训练机会来实现 职业目标候选人。