Genomic Analysis of Carbon Monoxide Utilization and Butanol Production by <em>Clostridium carboxidivorans</em> Strain P7^T

Increasing demand for the production of renewable fuels has recently generated a particular interest in microbial production of butanol. Anaerobic bacteria, such as Clostridium spp., can naturally convert carbohydrates into a variety of primary products, including alcohols like butanol. The genetics of microorganisms like Clostridium acetobutylicum have been well studied and their solvent-producing metabolic pathways characterized. In contrast, less is known about the genetics of Clostridium spp. capable of converting syngas or its individual components into solvents. In this study, the type of strain of a new solventogenic Clostridium species, C. carboxidivorans, was genetically characterized by genome sequencing. C. carboxidivorans strain P7T possessed a complete Wood-Ljungdahl pathway gene cluster, involving CO and CO2 fixation and conversion to acetyl-CoA. Moreover, with the exception of an acetone production pathway, all the genetic determinants of canonical ABE metabolic pathways for acetate, butyrate, ethanol and butanol production were present in the P7T chromosome. The functionality of these pathways was also confirmed by growth of P7T on CO and production of CO2 as well as volatile fatty acids (acetate and butyrate) and solvents (ethanol and butanol). P7T was also found to harbour a 19 Kbp plasmid, which did not include essential or butanol production related genes. This study has generated in depth knowledge of the P7T genome, which will be helpful in developing metabolic engineering strategies to improve C. carboxidivorans's natural capacity to produce potential biofuels from syngas.近年来，可再生燃料生产需求的持续攀升，使得微生物合成丁醇（butanol）的研究受到了广泛关注。厌氧菌（anaerobic bacteria）如梭菌属（Clostridium spp.），可自然将碳水化合物转化为多种初级产物，其中包含丁醇等醇类物质。丙酮丁醇梭菌（Clostridium acetobutylicum）这类微生物的遗传学特征已得到充分研究，其产溶剂的代谢通路也已被完整解析。与之相对，对于能够将合成气（syngas）或其单一组分转化为溶剂的梭菌属物种，其遗传学背景的研究仍较为匮乏。 本研究通过基因组测序技术，对新型产溶剂梭菌属物种——嗜羧基梭菌（C. carboxidivorans）的模式菌株P7^T进行了遗传学表征。该菌株携带完整的伍德-隆德哈尔途径（Wood-Ljungdahl pathway）基因簇，该通路涵盖一氧化碳（CO）与二氧化碳（CO2）的固定过程，并可将其转化为乙酰辅酶A（acetyl-CoA）。此外，除丙酮合成途径外，P7^T菌株的染色体上完整存在经典ABE代谢途径（ABE metabolic pathways）中与乙酸盐、丁酸盐、乙醇及丁醇合成相关的全部遗传决定因子。 上述代谢通路的功能亦得到了实验验证：P7^T菌株可在一氧化碳培养基中正常生长，并产生二氧化碳、挥发性脂肪酸（volatile fatty acids，包括乙酸与丁酸）以及溶剂（乙醇与丁醇）。研究同时发现，P7^T菌株携带一条19 kbp的质粒，该质粒未包含必需基因或丁醇合成相关的功能基因。 本研究深入解析了P7^T菌株的基因组信息，这将为开发代谢工程（metabolic engineering）策略提供理论支撑，以提升嗜羧基梭菌利用合成气生产潜在生物燃料（biofuels）的天然能力。