EAGER: Redox Regulation of Methanogenesis: A Basic Microbial Process at the Interface of Climate Change and Renewable Energy

智力优势：甲烷是一种重要的燃料，也是与全球变暖有关的主要温室气体。 目前自然界中甲烷的生物生产完全是由于产甲烷菌，一组严格厌氧的微生物，属于新认识的生命领域，古生菌。 通过在一个被称为产甲烷的过程中产生甲烷，这些生物促进了植物产生的纤维素和淀粉等复杂聚合物的厌氧降解，并在全球碳循环中发挥核心作用。 此外，其中二氧化碳和氢气转化为甲烷和水的甲烷生成模式之一被认为是地球上最古老的呼吸形式之一。 尽管人们对甲烷形成的生物化学途径了解很多，但对该过程如何响应环境条件的变化进行调节知之甚少。重要的是，目前还不知道产甲烷菌如何科普偶尔暴露于氧气的情况，这在它们的自然栖息地中很常见。 有可能，在适度的氧污染下，生物体改变了它们的细胞机制，使甲烷生成得以持续-这是一种重要的能力，因为甲烷的形成是它们唯一的能量来源。 初步研究表明，硫氧还蛋白（Trx）修饰的关键酶在这种控制策略中发挥作用。 Trx是一种普遍存在的调节蛋白，其与酶Trx还原酶合作，在代谢的精细控制中起作用。 然而，这一系统在产甲烷菌中仍然未被探索，尽管该组的每个成员都携带有编码这些蛋白质的潜力的基因。 在这个项目中，我们将测试的概念，Trx/Trx还原酶为基础的控制是一个决定因素的生物生产甲烷。 我们将利用两种产甲烷古菌作为模型：（一）詹氏甲烷球菌，一种根深蒂固的自养生物，栖息在深海热液喷口，只从二氧化碳和氢气中产生甲烷;（二）马氏甲烷八叠球菌，一种进化较晚的淡水产甲烷菌，具有更多样化的产甲烷能力。 将与基尔大学（德国）合作研究后一种微生物。 本研究的目的是：（i）鉴定M. jannaschii和M. Mazei使用产甲烷菌宿主中过度产生的蛋白质，从而提供与研究的体内相关性;（ii）鉴定Trx在这些生物体中修饰的酶和其他蛋白质;和（iii）确定基于Trx的修饰对选择的产甲烷酶的活性的影响。 这项工作将为更广泛的当代问题提供一个切入点，这些问题涉及自然栖息地中甲烷生成的控制，以及细菌、动物和植物的基于氧化还原的控制系统如何从起源于古细菌的控制系统进化而来。更广泛的影响：这项研究将阐明产甲烷微生物如何科普其栖息地暴露于氧气的挑战性影响。 因此，它将对能源生产、生物圈中碳的循环利用和全球变暖产生广泛的社会和科学影响。 更深入地了解甲烷形成的调节，提高了以有用的方式控制其生产的可能性？例如，在一个实施例中，在生物反应器和污水消化器中。该项目将通过培训两名研究生、四名本科生和一名高中生来整合研究和教育。 来自PI（“后基因组时代的微生物学”）指导的当前NSF-REU计划的本科生和高中教师将有机会参与这项研究。 将尽一切努力从各级任职人数不足的少数群体中征聘人员。 合作培训将拓宽弗吉尼亚理工大学、基尔大学和加州大学伯克利分校参与实验室的研究生的研究经验。 它的科学和社会相关性，加上不同职业阶段的各种调查人员的参与，将丰富该项目以及所有参与者的研究经验。

Intellectual merit: Methane is an important fuel and a major greenhouse gas linked to global warming. The current biological production of methane in nature is solely due to the methanogens a diverse group of strictly anaerobic microorganisms belonging to the newly recognized domain of life, the archaea. By generating methane in a process known as methanogenesis, these organisms facilitate the anaerobic degradation of complex polymers such as cellulose and starch that are produced by plants and, in so doing, play a central role in the global carbon cycle. Further, one of the modes of methanogenesis in which carbon dioxide and hydrogen are converted to methane and water is considered one of the most ancient forms of respiration on Earth. Although much has been learned about the biochemical pathways of methane formation, little is known as to how the process is regulated in response to change in environmental conditions. Importantly, it is not known how methanogens cope with sporadic exposure to oxygen, a common occurrence in their natural habitats. It is possible that, under moderate oxygen contamination, the organisms modify their cellular machinery so that methanogenesis is sustained--an important capability since methane formation represents their only source of energy. Preliminary studies indicate that modification of key enzymes by thioredoxin (Trx) plays a role in this control strategy. Trx is a ubiquitous regulatory protein that acts in partnership with an enzyme, Trx reductase, in the fine control of metabolism. However, this system remains unexplored in the methanogens, despite the fact that every member of this group carries genes with the potential for encoding these proteins. In this project we will test the concept that Trx/Trx reductase-based control is a determinant for the biological production of methane. We will utilize two methanogenic archaea as models: (i) Methanocaldococcus jannaschii, a deeply rooted autotroph that inhabits deep-sea hydrothermal vents and produces methane exclusively from carbon dioxide and hydrogen, and (ii) Methanosarcina mazei, a late-evolving freshwater methanogen with more diverse methanogenic capability. The latter organism will be investigated in collaboration with the University of Kiel (Germany). The objectives of the study are: (i) To characterize the putative Trxs and Trx reductases of M. jannaschii and M. mazei using proteins over-produced in a methanogen host, thereby providing in vivo relevance to the investigation; (ii) To identify the enzymes and other proteins that Trx modifies in these organisms; and (iii) To determine the effect of Trx-based modification on the activity of select methanogenic enzymes. The work will provide an entrée to broader contemporary questions relating to the control of methanogenesis in natural habitats and to how redox-based control systems of bacteria, animals and plants evolved from those originating in the archaea.Broader impact: This research will elucidate how methane-producing microorganisms cope with the challenging effects of exposure to oxygen in their habitats. Consequently, it will have broad societal and scientific implications relating to energy production, recycling of carbon in the biosphere and global warming. Greater insight into the regulation of methane formation raises the possibility of controlling its production in a useful manner?e.g., in bioreactors and sewage digesters. The project will integrate research and education through the training of two graduate students, four undergraduates, and a high school student. Undergraduate students and high school teachers from a current NSF-REU program directed by the PI ("Microbiology in the Post-Genome Era") will have an opportunity to participate in this research. Every effort will be made to recruit from underrepresented minority groups at all levels. The collaborative training will broaden the research experience of graduate students in the participating laboratories at Virginia Tech, University of Kiel and University of California-Berkeley. Its scientific and societal relevance coupled with the participation of a diverse group of investigators at different career stages will enrich the project as well as the research experience of all participants.