Nitric oxide reactions in metalloenzymes

金属酶中的一氧化氮反应

• 批准号: 8292380
• 负责人: PIERRE MOENNE-LOCCOZ
• 金额: $ 27.87万
• 依托单位: OREGON HEALTH & SCIENCE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2006
• 资助国家: 美国
• 起止时间: 2006-04-01 至 2016-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8292380
• 关键词: Active Sites; Archaea; Bacteria; Binding; Biochemical Reaction; Biomedical Engineering; Catalysis; Characteristics; Chemistry; Complex; Copper; Coupling; Drug Metabolic Detoxication; Electron Spin Resonance Spectroscopy; Electron Transport; Engineering; Environment; Enzymes; Event; Exhibits; Family; Flavin Mononucleotide; Freezing; Goals; Heme; Hemerythrin; Human; Hydrogen Peroxide; Immune response; Ions; Iron; Kinetics; Lead; Modeling; Molecular; Monitor; Myoglobin; Nitric Oxide; Noble Gases; Orthologous Gene; Oxidases; Oxidoreductase; Pharmaceutical Preparations; Play; Process; Proteins; Protons; Raman Spectrum Analysis; Reaction; Research; Resistance; Role; Route; Site; Spectroscopy, Fourier Transform Infrared; Spectrum Analysis; Structure; System; Variant; X-Ray Crystallography; bacterial resistance; carboxylate; cofactor; cold temperature; combat; enzyme model; heme a; hyponitrite; insight; iron nitrosyl; metalloenzyme; microbial; microorganism; nitroxyl; pathogen; photolysis; protonation; public health relevance; research study

DESCRIPTION (provided by applicant): The long-term goal of this research is to elucidate the reaction mechanisms of NO reduction that occur at heme and non-heme diiron centers of bacterial metalloenzymes. Our studies will focus on two enzymatic systems and related bioengineered or synthetic models: 1) NO-detoxifying flavodiiron proteins (FDPs) and 2) denitrifying NO reductases (denNORs). All of the proteins within these two families are known to reduce NO to the unreactive N2O product, but they do so with wide variation in efficiency and protein matrix structure. Although several of these proteins have been characterized by X-ray crystallography, the initial steps of NO binding, iron-nitrosyl reduction, and how these catalytic events differ between systems are not well understood. The coupling of resonance Raman, FTIR, and EPR spectroscopies with rapid-freeze-quench analyses provides unique capabilities to define NO-binding geometries at diiron clusters and to follow the N-N bond formation, N-O bond cleavage, and protonation steps that must take place to convert two NO molecules to N2O and H2O. Studying a diverse group of native enzymes and models will allow us to compare and contrast structural information on iron-nitrosyl intermediates and the efficiency of the reductive and proton transfer steps of this reaction. Public health relevance: A better understanding of microbial NO reductases is highly desirable since these enzymatic reactions lead to microorganisms' resistance to the mammalian immune response. Furthermore, there are no human orthologs to these microbial enzymes; they represent potential targets for new drugs. PUBLIC HEALTH RELEVANCE: The goal of this research is to elucidate the mechanisms of NO detoxification employed by microorganisms to combat the mammalian immune response. While many metalloenzymes involved in this process are structurally distinct from one another, they have in common an active site containing two iron ions. To understand the reaction of these diiron sites with NO will require a combination of rapid kinetic analyses and complementary molecular spectroscopies.

描述（由申请人提供）：本研究的长期目标是阐明细菌金属酶的血红素和非血红素二铁中心发生的NO还原反应机制。我们的研究将集中在两个酶系统和相关的生物工程或合成模型：1）NO解毒黄二铁蛋白（FDP）和2）还原NO还原酶（denNORs）。已知这两个家族中的所有蛋白质都能将NO还原为非反应性N2 O产物，但它们的效率和蛋白质基质结构存在很大差异。虽然这些蛋白质中的几个已经通过X射线晶体学表征，但NO结合、铁-亚硝酰还原的初始步骤以及这些催化事件在系统之间的差异还不清楚。共振拉曼，FTIR和EPR光谱与快速冷冻淬火分析的耦合提供了独特的能力，以定义NO-结合的几何形状在二铁簇，并遵循N-N键形成，N-O键断裂，和质子化的步骤，必须发生两个NO分子转化为N2 O和H2O。研究一组不同的天然酶和模型将使我们能够比较和对比铁-亚硝基中间体的结构信息和该反应的还原和质子转移步骤的效率。公共卫生相关性：更好地了解微生物NO还原酶是非常需要的，因为这些酶促反应导致微生物对哺乳动物免疫应答的抗性。此外，这些微生物酶没有人类直系同源物;它们代表了新药的潜在靶点。 公共卫生相关性：本研究的目的是阐明微生物利用NO解毒来对抗哺乳动物免疫反应的机制。虽然许多参与这一过程的金属酶在结构上彼此不同，但它们都有一个含有两个铁离子的活性位点。要了解这些二铁网站与NO的反应将需要快速动力学分析和互补的分子光谱学相结合。