Particulate Methane Monooxygenase

颗粒甲烷单加氧酶

• 批准号: 7942225
• 负责人: AMY C. ROSENZWEIG
• 金额: $ 5.1万
• 依托单位: NORTHWESTERN UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2009
• 资助国家: 美国
• 起止时间: 2009-09-30 至 2010-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7942225
• 关键词: Active Sites; Affinity; Ammonia; Architecture; Arts; Bacteria; Bathing; Binding; Binding Sites; Bioinorganic Chemistry; Biological Assay; Bioremediations; Carbon; Carcinogens; Chemistry; Copper; Crystallization; Data; Energy-Generating Resources; Enzymes; Escherichia coli; Essential Amino Acids; Fab Immunoglobulins; Freezing; Gases; Global Warming; Goals; Health; Human; Hydrocarbons; In Vitro; Knowledge; Ligands; Lipids; Location; Membrane; Membrane Proteins; Metabolic Pathway; Metals; Methane; Methane hydroxylase; Methanol; Methods; Methylococcus capsulatus; Methylocystis; Methylosinus trichosporium; Microfluidics; Nature; Nitrosomonas; Organism; Particulate; Phage Display; Play; Preparation; Procedures; Proteins; Raman Spectrum Analysis; Research; Research Personnel; Resolution; Rhodobacter sphaeroides; Role; Sampling; Screening procedure; Site-Directed Mutagenesis; Soil; Structure; Substrate Specificity; System; Techniques; Testing; Transmembrane Domain; Variant; Water; Work; ammonia monooxygenase; analog; carboxylate; catalyst; enzyme activity; improved; oxidation; programs; protein complex; public health relevance; reconstitution; research study

DESCRIPTION (provided by applicant): Methanotrophic bacteria utilize methane as their sole carbon and energy source. The first step in their metabolic pathway is the oxidation of methane to methanol by methane monooxygenase (MMO) enzyme systems. All but one genus of methanotrophs produce a membrane-bound, copper-containing enzyme called particulate methane monooxygenase (pMMO). Although pMMO is the predominant methane oxidation catalyst in nature, it has proved difficult to isolate, and most investigators have instead opted to study soluble methane monooxygenase (sMMO), a diiron carboxylate-bridged enzyme that is more tractable, but less universal, than pMMO. The structure and mechanism of pMMO and the homologous enzyme ammonia monooxygenase (AMO) remain one of the major unsolved problems in bioinorganic chemistry. Understanding how pMMO activates O2 for oxidation of methane and other hydrocarbons is the long term goal of this research program. Despite the availability of a crystal structure and extensive spectroscopic data, key questions regarding the metal content and active site identity remain unanswered. These issues are of fundamental importance to bioinorganic copper chemistry and have implications for the use of methanotrophs in bioremediation. In addition, methanotrophs play a key role in the global carbon cycle and could help mitigate the deleterious effects of global warming on human health. The proposed research involves purification and characterization of pMMO and AMO from multiple organisms. State-of-the-art crystallization techniques for membrane proteins will be applied to these enzymes. In addition, expression systems will be developed to enable site-directed mutagenesis experiments. Finally, in vitro enzyme activity will be optimized and mechanistic studies initiated. PUBLIC HEALTH RELEVANCE: Bacteria that consume methane gas play an important role in mitigating global warming, which has deleterious effects on human health. These bacteria also are useful for bioremediation of soil and water polluted with hydrocarbon carcinogens. This project will investigate the details of how these bacteria transform methane into methanol.

描述（由申请人提供）：甲烷营养细菌利用甲烷作为其唯一的碳和能源。其代谢途径的第一步是通过甲烷单加氧酶（MMO）酶系统将甲烷氧化为甲醇。除一种甲烷营养的属外，所有属都会产生一种膜结合的，含铜的酶，称为颗粒甲烷单加氧酶（PMMO）。尽管PMMO本质上是主要的甲烷氧化催化剂，但事实证明它很难分离，而大多数研究者则选择研究一种可溶性甲烷单加氧酶（SMMO），这是一种甲羧酸酯桥桥的酶，它是可拖动的，但比PMMO较少，但较少。 PMMO的结构和机制和同源酶单加氧酶（AMO）仍然是生物无机化学中主要未解决的问题之一。了解PMMO如何激活O2来氧化甲烷和其他烃是该研究计划的长期目标。尽管有晶体结构和广泛的光谱数据，但有关金属含量和主动位点标识的关键问题仍未得到解答。这些问题对生物有机铜化学具有​​至关重要的重要性，并且对在生物修复中使用甲嗜性具有影响。此外，甲烷营养在全球碳周期中起着关键作用，并有助于减轻全球变暖对人类健康的有害影响。拟议的研究涉及多种生物的PMMO和AMO的纯化和表征。膜蛋白的最先进的结晶技术将应用于这些酶。此外，将开发表达系统以实现位置定向的诱变实验。最后，将优化体外酶活性并开始机械研究。公共卫生相关性：消耗甲烷气体的细菌在减轻全球变暖方面起着重要作用，这对人类健康产生了有害影响。这些细菌也可用于对土壤和用碳氢化合物致癌物污染的水进行生物修复。该项目将研究这些细菌如何将甲烷转化为甲醇的细节。