Interspecies Metabolic Complementation in Geothermal Microbial Mats

这项研究将确定自然微生物生态系统中负责一氧化碳厌氧氧化和相关代谢(例如，氢气形成)的基因并对其进行排序。细菌和古生菌从非常广泛的来源获得能量和碳。一些微生物可以利用一氧化碳(对人类剧毒)作为能源和碳积木来生产新的生物质和细胞。这些微生物中的一部分会产生与消耗一氧化碳有关的氢气。几乎没有分离出能产生氢气的羧基营养细胞，但很可能有许多尚未培养的微生物具有编码这种代谢能力的基因。在这项研究中，SIGEX(底物诱导基因表达--一种新的分子生物学技术)将被用来从偏远地区堪察加乌松火山口的自然微生物种群中寻找这些基因。大量DNA将从自然和实验室培养的微生物垫和温泉环境中的沉积物中提取。然后，DNA将被切成短片段，克隆到大肠杆菌中的分子报告结构中，并筛选是否存在感兴趣的基因。这一战略将基于研究人员最近从黄石和西伯利亚温泉获得的羧基营养细胞的基因组序列。在一氧化碳存在的情况下启动的基因将被测序，并将评估具有这种代谢活动的微生物的多样性。许多产氢基因与一氧化碳氧化基因密切相关，并将通过这种筛选方法进行跟踪。环境化学将通过使用微电极和扫描伏安法来测量沐浴在微生物垫上的水的化学成分，从而将环境化学与基因组潜力联系起来。这项工作将是利用SIGEX寻找稀有但在环境和生物技术上具有重要意义的基因的首批研究之一。它还将拓宽我们对微生物一氧化碳氧化产氢的理解。对这些基因的检测和对这些基因编码的酶的检测，最终可能会为开发出模仿微生物氧化一氧化碳和产生氢的效率的工程生物催化剂铺平道路。如果像研究人员假设的那样，一氧化碳敏感基因被发现广泛存在，这可能表明一氧化碳是微生物生态系统中碳循环中非常活跃的组成部分。这项研究将支持研究生(具有分子生物学背景)和博士后研究员(具有地球化学背景)的跨学科培训。这项工作将通过与旧金山探索博物馆的合作向普通公众传达，旧金山探索博物馆是世界著名的科学中心，将艺术、科学和人类感知联系在一起。

This research will identify and sequence the genes responsible for anaerobic oxidation of carbon monoxide and related metabolism (e.g., hydrogen formation) within natural microbial ecosystems. Bacteria and Archaea obtain their energy and carbon from a very wide variety of sources. Some microbes can use carbon monoxide (highly toxic to humans) as both an energy source and a carbon building block for producing new biomass and cells. A subset of these microbes produces hydrogen gas in connection with consuming carbon monoxide. Very few hydrogen-producing carboxydotrophs have been isolated, but there are likely many microbes not yet cultured that have genes that encode for this metabolic capability. In this research, SIGEX (Substrate Induced Gene Expression -- a novel molecular biology technique) will be used to hunt down these genes from natural populations of microbes in a remote site, the Uzon Caldera, Kamchatka. Bulk DNA will be extracted from natural and laboratory cultures of microbial mats and sediments from hot spring environments. The DNA will then be cut into short pieces, cloned into a molecular reporter construct in E. coli, and screened for presence or absence of the genes of interest. This strategy will be based on the investigators' recently obtained genome sequence of carboxydotrophs from Yellowstone and Siberian hot springs. Genes that switch on in the presence of carbon monoxide will be sequenced, and the diversity of microbes with this metabolic activity will be assessed. Many of the genes for hydrogen production are closely associated with carbon monoxide oxidation genes and will track with this screening method. Environmental chemistry will be linked to genomic potential by using microelectrodes and scanning voltammetry to measure the chemical composition of the water bathing the microbial mats. This work will be one of the first studies to employ SIGEX for finding rare but environmentally and biotechnologically important genes. It will also broaden our understanding of hydrogen production through microbial carbon monoxide oxidation. The detection of such genes and examination of the enzymes encoded by these genes may eventually pave the way for engineered biocatalysts that would mimic the efficiency of the microbes in oxidizing carbon monoxide and producing hydrogen. If carbon monoxide sensitive genes are found to be widespread, as the investigators' hypothesis states, it could indicate that carbon monoxide is a very active component of the carbon cycle in microbial ecosystems. This research will support the interdisciplinary training of a graduate student (with a molecular biology background) and a postdoctoral fellow (with a geochemistry background). The work will be conveyed to the general public through a partnership with the San Francisco Exploratorium, a world-renowned science center that links art, science, and human perception.