Developing Methanosarcina spp. as a model system to study cytochromes c and their role in archaeal methane metabolism

正在开发甲烷八叠球菌属。

• 批准号: 10679362
• 负责人: Dinesh Gupta
• 金额: $ 6.95万
• 依托单位: UNIVERSITY OF CALIFORNIA BERKELEY
• 依托单位国家: 美国
• 财政年份: 2023
• 资助国家: 美国
• 起止时间: 2023-09-30 至 2025-09-29
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10679362
• 关键词: Anaerobic Bacteria; Archaea; Atmosphere; Binding; Biochemical; Biogenesis; Biological Models; Biotechnology; CRISPR/Cas technology; Coculture Techniques; Complex; Consumption; Coupled; Development; Electron Transport; Electrons; Engineering; Environment; Escherichia coli; Evaluation; Ferredoxin; Gene Expression; Genes; Genetic; Genetic Models; Genome; Genomics; Goals; Growth; Health; Heme; Human; In Vitro; Knowledge; Mediating; Membrane; Metabolic; Methane; Methane Metabolism Pathway; Methanobacteria; Methanosarcina; Methanosarcina barkeri; Modeling; Molecular; NADH; Nature; Nitrogen Fixation; Operon; Organism; Oxidation-Reduction; Pathway interactions; Physiological; Play; Postdoctoral Fellow; Process; Production; Proteins; Reaction; Research; Rhodobacter; Role; Source; Symbiosis; System; Techniques; biochemical tools; citrate carrier; climate change; climate crisis; cofactor; cytochrome c; genome editing; greenhouse gases; improved; in vivo; insight; interest; methanophenazine; microorganism; oxidation; reconstitution; sodium ion; success; sulfate reducing bacteria; technology development; tool

Project Summary/Abstract Cytochromes c are crucial in methane-metabolizing archaea for the production and consumption of methane coupled to growth and energy conservation. Overall, methane-metabolizing archaea mediate the net flux of methane released into the atmosphere and thus, significantly impact the global methane cycle and climate change. Based on genomic studies, Cytochromes c have been hypothesized to play an important role in methane metabolism however the underlying molecular mechanisms remain elusive, primarily due to the absence of a well-developed genetic model system. Even attempts to study archaeal cytochrome c using an alternative approach such as heterologous expression in well-established bacterial systems have not proven successful so far. The main focus of the proposed research is to develop a genetically tractable methanogenic archaeon, Methanosarcina acetivorans, as a platform to functionally characterize archaeal cytochrome c and gain physiological insights into the role of cytochrome c in methane metabolism across different archaeal species. Cytochromes c are ubiquitous electron transfer proteins that require a covalent attachment to its heme co-factor, a process called cytochrome c biogenesis. Coordination between the cytochrome c biogenesis pathway and the cytochrome c of interest is critical for the successful production of a functional cytochrome c in a heterologous host. Using genetic and biochemical tools, I have recently characterized the cytochrome c biogenesis pathway in the model methanogenic archaeon, Methanosarcina acetivorans. Using this knowledge, the project aims to develop M. acetivorans as a genetic chassis to produce and functionally characterize archaeal cytochrome c from diverse archaeal species. Aim1 of the project will functionally characterize the crucial cytochromes c belonging to methane-producing archaea or methanogens, and Aim2 will study these proteins from methane-consuming archaea using both in vitro and in vivo analyses. This research will improve our understanding of methane metabolism and lead to the development of an archaeal host to study cytochrome c proteins from archaea. Ultimately, the knowledge garnered from these studies can be used to develop sustainable solutions for the global climate crisis and mitigate its harmful impacts on human health.

项目摘要/摘要 细胞色素C在甲烷 - 代谢古细菌中至关重要，用于生产和消耗甲烷 与生长和节能相结合。总体而言，甲烷 - 代谢的古细菌介导了净通量 甲烷释放到大气中，从而显着影响全球甲烷周期和气候 改变。基于基因组研究，已经假设细胞色素C在 甲烷代谢然而，潜在的分子机制仍然难以捉摸，主要是由于 缺乏发达的遗传模型系统。甚至尝试使用一个 尚未证明替代方法，例如良好的细菌系统中的异源表达 到目前为止成功。拟议的研究的主要重点是开发一种可遗传障碍的甲烷作用 甲壳虫乙烯菌的古老，是一个功能表征古细菌C和的平台 对细胞色素c在不同古细菌的甲烷代谢中的作用获得生理见解 物种。 细胞色素C是无处不在的电子转移蛋白，需要在其血红素co因子上共价附着 一个称为细胞色素C生物发生的过程。细胞色素C生物发生途径与 感兴趣的细胞色素C对于在异源中成功产生功能性细胞色素C至关重要 主持人。使用遗传和生化工具，我最近表征了细胞色素C生物发生途径 在模型甲烷元素的甲烷菌乙酰乙烷中。使用这些知识，该项目的目的是 开发乙酰维多氏菌作为遗传底盘，以产生和功能表征古细胞色素c 来自多种古细菌。该项目的AIM1将在功能上表征至关重要 属于产生甲烷的古细菌或甲烷的细胞色素c，AIM2将研究这些蛋白质 使用体外和体内分析的甲烷消费古细菌。这项研究将改善我们的 了解甲烷代谢并导致古细菌宿主的发展以研究细胞色素c 古细菌的蛋白质。最终，从这些研究中获得的知识可用于发展 全球气候危机的可持续解决方案，并减轻其对人类健康的有害影响。