Proton-Coupling Strategies in Alkaliphilic Bacillus

嗜碱芽孢杆菌中的质子耦合策略

• 批准号: 7939051
• 负责人: Terry Ann Krulwich
• 金额: $ 8.48万
• 依托单位: ICAHN SCHOOL OF MEDICINE AT MOUNT SINAI
• 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/7939051
• 关键词: ATP Synthesis Pathway; Accounting; Acids; Alkalies; Antibiotics; Area; Bacillus (bacterium); Bacteria; Biochemical Genetics; Bioenergetics; Biological Assay; Biological Models; Carrier Proteins; Cell Respiration; Cells; Chemicals; Cholates; Complement; Complex; Data; Development; Elements; Gene Proteins; Genomics; Goals; Growth; Individual; Ion Cotransport; Ions; Membrane; Mitochondria; Molecular; Movement; Operon; Oxidases; Oxidative Phosphorylation; Oxygen; Pathway interactions; Pattern; Physiological; Play; Process; Production; Property; Proteins; Protons; Research Personnel; Respiration; Respiratory Chain; Role; Site; Sodium-Hydrogen Antiporter; Solutions; Structure; System; Testing; Treatment Protocols; Vesicle; Work; antiport; antiporter; base; cytochrome c oxidase; driving force; electron donor; inhibitor/antagonist; insight; mutant; novel; pH Homeostasis; pH gradient; pathogen; programs; protein protein interaction; research study; stoichiometry; uptake

DESCRIPTION (provided by applicant): This project will clarify the mechanisms used by alkaliphilic Bacillus for the two processes that move protons inward at high external pH: how extremely robust proton-coupled oxidative phosphorylation (OXPHOS) occurs under conditions in which the chemiosmotic driving force (negative and alkaline in) is very low; and how the novel properties of the Mrp sodium/proton antiporter underpin its capacity to support remarkable alkaline pH homeostasis, resulting in a "reversed pH gradient" at external pH values such that the cytoplasmic pH is more than 2 units below an optimal external pH of 10.5. Three specific aims will focus on the following questions, (i) How do specific features of the membrane-embedded a- and c-subunits of ATP synthase make distinct critical contributions to proton capture and movement through the ATP synthase? The approaches will include analyses of site-directed mutants that alter properties of critical ATP synthase residues in the proton path and studies of the c-subunit stoichiometry of the synthase. (ii) What is the special involvement of a particular respiratory chain complex, the Cta cytochrome oxidase, in partnering with the ATP synthase at high pH for sequestered proton transfers to the synthase? Protein-protein interactions between Cta and ATP synthase as well as specific conditions and features of the two complexes involved in such interactions will be the focus, (iii) What are the roles of the seven Mrp proteins and the properties of the Mrp antiporter that account for its unusual complexity and efficacy? A multi-pronged biochemical, genetic and physiological approach will test the hypotheses that: (a) this secondary sodium/proton antiporter functions as a novel, hetero-oligomeric complex; and (b) some Mrp proteins have distinct catalytic activities that have positive synergy with Mrp-dependent antiport. Two areas are studied bacteria that thrive in alkali. They are a model system for unraveling details of chemical energy production (as ATP) by oxygen-dependent cells. These bacteria are hypothesized to share strategies with mitochondria so the findings on ATP synthesis have pharmacalogical implications for normal and pathological states of cellular respiration. The studies of the Mrp system characterize a novel protein carrier found in many bacterial pathogens. Mrp is a potential target for development of new antibiotics.

描述（由申请人提供）：该项目将阐明碱性芽孢杆菌在高外部pH下向内移动质子的两个过程的机制：在化学驱动力（负驱动力和碱性）下，在高外部pH中向内移动质子是多么强大的质子偶联氧化磷酸化（OXPHOS）。以及MRP钠/质子抗腐蚀剂的新特性如何支持其支持显着的碱性pH稳态的能力，从而在外部pH值处产生“反向的pH梯度”，从而使细胞质pH值超过2个单位，低于最佳外部pH值10.5。 （i）ATP合酶的膜包裹的A和C-亚基的特定特征如何为质子捕获和通过ATP合酶运动做出明显的关键贡献？这些方法将包括对位置定向突变体的分析，这些突变体改变了质子路径中关键ATP合酶残基的性质以及合成酶C-亚基化学计量的研究。 （ii）特定呼吸链复合物（CTA细胞色素氧化酶）与高pH值的ATP合酶合作以隔离质子转移到合成酶时，特定的呼吸链复合物的特殊参与是什么？ CTA和ATP合酶之间的蛋白质 - 蛋白质相互作用以及这种相互作用中涉及的两个复合物的特定条件和特征将是重点，（iii）七种MRP蛋白的作用以及MRP抗毒剂的特性，其占其异常复杂性和功效的MRP抗植物特性是什么？一种多管齐的生化，遗传和生理方法将检验：（a）这种次级钠/质子抗胞剂作为一种新颖的，异性 - 寡聚体复合物； （b）某些MRP蛋白具有不同的催化活性，与MRP依赖性的抗移植物具有正协同作用。 研究了两个区域在碱中壮成长的细菌。它们是一个模型系统，可通过氧依赖性细胞揭示化学能产生（ATP）的细节。假设这些细菌可以与线粒体共享策略，因此ATP合成的发现对正常和病理呼吸的病理状态具有药物学意义。 MRP系统的研究表征了在许多细菌病原体中发现的新型蛋白质载体。 MRP是开发新抗生素的潜在靶标。