Nitric Oxide Reactions in Metalloenzymes

金属酶中的一氧化氮反应

基本信息

批准号：
7385993
负责人：
PIERRE MOENNE-LOCCOZ
金额：
$ 21.97万
依托单位：
OREGON HEALTH & SCIENCE UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
2006
资助国家：
美国
起止时间：
2006-04-01 至 2011-03-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/7385993
关键词：
Azides Bacillus (bacterium)Bacteria Binding Biochemical Reaction Catalytic Domain Chemicals Complex Copper Diffuse Distal Docking Electron Transport Electrons Environment Enzymes Event Fees Film Flavoproteins Freezing Frequencies Gases Goals Health Heme Heme Iron Hemeproteins Human Hydrogen Bonding Immune response Ions Iron Label Location Maps Measures Metals Methodology Methods Modeling Nitric Oxide Numbers Object Attachment Oxidases Oxidoreductase Paracoccus denitrificans Performance Proteins Protons Reaction Research Resistance Rest Role Route Side Site Spectroscopy, Fourier Transform Infrared Structure Structure-Activity Relationship Testing Thermus thermophilus Thinking Variant Water analog cold temperature cytochrome ba3 dimer electron density enzyme structure enzyme substrate heme a3 hyponitrite insight instrumentation iron nitrosyl metalloenzyme microbial novel pathogenic bacteria photolysis protonation research study

项目摘要

DESCRIPTION (provided by applicant): This research aims to elucidate reaction intermediates of NO-reductase activity in diiron proteins, with the ultimate goal of understanding how the metal clusters catalyze this reaction. Our studies will focus on three enzymes: 1) denitrifying NO reductases cNOR from Paracoccus denitrificans and qCuANOR from Bacillus azotoformans, 2) the [heme-copper] ba3 terminal oxidase from Thermus thermophilus, and 3) detoxifying NO reductase flavoprotein A (FprA) from Moorella thermoacetica. A better understanding of microbial NO reductases is highly desirable considering that these enzymatic reactions provide a resistance to the mammalian immune response. Although crystal structures exist for some of these enzymes, the structure and reactivity of their NO-complexes are not known. Diiron proteins participate in both detoxifying and denitrifying NO reductase reactions and are thought to react with NO to form [FeNO]2 intermediates. Alternative mechanistic models are considered and tested in this proposal. Resonance Raman and FTIR spectroscopies have the unique capability to identify nitrosyl intermediates and to define their NO-binding geometries with regard to the two metal ions. Novel rapid freeze-quench (RFQ) instrumentation that can trap intermediates within a sub-ms timescale provides new opportunities to characterize reaction intermediates that were previously inaccessible to spectroscopic methods. FTIR spectroscopy, in conjunction with low temperature photolysis of N3~ and CO, acting as NO surrogates, offers insight into binding geometries, potential hydrogen bond interactions and proton transfers relevant to these NO reductase mechanisms.

描述（由申请人提供）：这项研究旨在阐明二铁蛋白中NO-还原酶活性的反应中间体，其最终目标是了解金属簇如何催化这种反应。我们的研究将侧重于三种酶：1）硝化酶denitrificans和Qcuanor cnor cnor conor和Qcuanor，来自氮杂杆菌芽孢杆菌，2）[heme-copper] ba3末端氧化酶来自Thermus hyterus hyterophilus，and thermus hythophilus和3）从Moorealla（Fravopra）中脱氧氧化氧化物（3）取消。考虑到这些酶促反应对哺乳动物免疫反应具有抗性，对微生物无还原酶的更好理解是非常可取的。尽管其中一些酶存在晶体结构，但其无复合物的结构和反应性尚不清楚。二铁蛋白参与排毒和反硝化无还原酶反应，并被认为反应不形成[Feno] 2个中间体。在此提案中考虑并测试了替代机械模型。共振拉曼和FTIR光谱具有独特的能力，可以识别亚硝基中间体，并针对两个金属离子定义其无结合的几何形状。可以将中间体捕获在子-MS时段中的新型快速冻结仪（RFQ）仪器为表征谱系方法无法访问的反应中间体提供了新的机会。 FTIR光谱与N3〜和CO的低温光解相结合，充当不替代物，可深入了解结合几何形状，潜在的氢键相互作用以及与这些无还原酶机制相关的质子转移。