Project 5: Molecular Insight into Dioxin Degradation by Microbes and Microbial

项目摘要（参见说明）： 微生物世界经过 37 亿年的进化而呈现出巨大的多样性，这既提供了未被发现的代谢能力（包括污染物降解能力）的机会，也提供了检测、恢复和表征这种活动的挑战。虽然几千年来，由于森林火灾等自然事件，微生物一直接触到少量的二恶英，但直到上个世纪左右，二恶英才因人类活动而成为自然界丰富的食物来源。这可以解释为什么只有少数特征明确的微生物菌株具有矿化二恶英的能力。我们建议描述纯培养物和微生物群落中微生物对二恶英的反应，以了解此类危险化合物的环境解毒的局限性。我们的长期目标是开发用于增强生物降解的微生物，识别分子标记以帮助现场评估，并开发预测污染场地生物降解的模型。我们在本提案中的具体目标是：(1) 阐明氯化二恶英降解的生理学、生物化学和遗传学，以便了解和改善这些剧毒污染物的修复；(2) 通过对微生物和活性微生物群落进行生理基因组学研究来评估（元）基因组范围内的反应和代谢能力，以响应氯化二恶英，包括来自环境重要的地质吸附剂的氯化二恶英；(3) 探索和回收大自然的催化多样性，目标是开发微生物群落代谢能力的全面概况，以降解和/或矿化环境中的氯化二恶英和二恶英类化合物。我们建议使用富集来帮助识别功能活跃的群体和基因，使用稳定同位素探测从活跃的降解群体中回收DNA，使用宏基因组和宏转录组文库以及目标DNA和RNA的大规模平行RNA诱饵捕获的新技术来恢复完整的基因和操纵子，使用这些信息来了解二恶英降解的生物化学，并使用二恶英降解模型细菌的转录组和生化分析来了解生物体对降解过程中涉及的各种压力（即底物和代谢物毒性、代谢通量等）的总反应。

PROJECT SUMMARY (See instructions): The microbial world is hugely diverse owing to its 3.7 billion years of evolution, which provides for both the opportunity of undiscovered metabolic capacity, including that for pollutant degradation, and the challenge of detecting, recovering, and characterizing this activity. While microbes have been exposed to small amounts of dioxins over millennia due to natural events such as forest fires, it is only in the last century or so that dioxins have become an abundant food source in nature due to anthropogenic activity. This may explain why there are only a few well characterized microbial strains with the ability to mineralize dioxins. We propose to characterize the microbial response to dioxins in pure cultures and microbial communities to understand the limitations on environmental detoxification of this hazardous class of compounds. Our long term goal is to develop microbes for enhanced biodegradation, to identify molecular markers to aid in site assessment, and to develop models for prediction of biodegradation at contaminated sites. Our Specific Aims in the present proposal are: (1) to elucidate the physiology, biochemistry, and genetics of chlorinated dioxins degradation in order to understand and improve the remediation of these highly toxic pollutants, (2) to assess (meta)genome-wide responses and metabolic capabilities by conducting physiogenomic studies of microorganisms and active microbial communities in response to chlorinated dioxins including those from environmentally important geosorbents, and (3) to explore and recover nature's catalytic diversity with the goal of developing a comprehensive profile of microbial community metabolic capabilities for degradation and/or mineralization of chlorinated dioxins and dioxin-like compounds in the environment. We propose to use enrichments to help identify the functionally active populations and hence genes, to use stable isotope probing to recover DNA from the active degrading populations, to use metagenomic and metatranscriptomic libraries and the novel technology of massive parallel RNA bait capture of target DNA and RNA to recover the full genes and operons, to use this information to understand the biochemistry of dioxins degradation, and to use transcriptomic and biochemical analyses of a model dioxin degrading bacterium to understand the total response of the organism to the various pressures involved during the degradation process (i.e. substrate and metabolite toxicity, metabolic fluxes, etc.).