Eliciting novel microbial phenotypes through transcriptional, degradation, and translational engineering

生物技术项目化学、生物工程、环境和运输系统分部通过转录、降解和转化工程筛选新的微生物表型gregory stephanopoulos麻省理工学院这项研究的目标是选择转录和翻译机制的成员，以丰富全球监管机构的库，通过这些监管机构，新的，否则无法到达的，表型可以在原核细胞中引起。量化各种调控因子及其组合工程产生的文库多样性的方法将作为指导最有前途的候选调控因子的表型改进过程的手段。最后，一系列专门设计的实验将阐明突变调节因子与宿主细胞的天然机制相互作用以引发新表型的机制。这项研究的智力价值在于促进了对转录机制及其与全球细胞内到达的突变调节因子的相互作用的基本理解。此外，它将阐明转录调控通过识别直接和间接的分子相互作用，基因表达受到影响。也许最持久的影响将是认识到许多细胞表型是许多基因的集体属性，因此，应该通过比单基因调节更有创造性的方法来优化，如本文提出的全球调节工程。该研究的更广泛的影响是打开了一个新的，完全未开发的，途径的表型启发和品系改进。当考虑到对有毒化合物的耐受性的特定表型时，这是特别重要的。作为一个模型系统，大肠杆菌对乙醇和其他有毒生物质水解副产物的耐受性将被研究。通过设计更具耐受性的菌株，整个生物质到生物燃料的转化过程将大大改善。目前因耐受性有限而受损的其他生物技术过程也可能同样受益于这项研究。

Biotechnology ProgramDivision of Chemical, Bioengineering, Environmental, and Transport SystemsEliciting Novel Microbial Phenotypes through Transcriptional, Degradation, and Translational EngineeringGregory StephanopoulosMassachusetts Institute of TechnologyCBET-0730238The goal of this research is to select members of the transcription and translation machinery in order to enrich the repertoire of global regulators by which new, otherwise unreachable, phenotypes can be elicited in prokaryotic cells. Methods to quantify the diversity of libraries resulting from the engineering of various regulators and their combinations will be developed as means of guiding the phenotype improvement process to the most promising candidate regulators. Finally, a series of specially designed experiments will elucidate the mechanism by which mutant regulators interact with the native machinery of the host cell in eliciting the new phenotypes. The intellectual merit of this research lies in advancing the fundamental understanding of the transcription mechanism and its interaction with mutant regulators of global intracellular reach. Additionally, it will elucidate transcriptional regulation by identifying direct and indirect molecular interactions by which gene expression is affected. Perhaps the most lasting impact will be the realization that many cellular phenotypes are the collective property of many genes and, as such, should be optimized by more creative methods than single gene modulation, like global regulator engineering proposed herein. The broader impact of the research is the opening of a new, totally unexplored, avenue to phenotype elicitation and strain improvement. This is of particular importance when one considers the specific phenotypes of tolerance to toxic compounds. As a model system, tolerance of E. coli to ethanol and other toxic byproducts of biomass hydrolysis will be studied. By engineering more tolerant strains, an overall biomass-to-biofuels conversion process will be substantially improved. Other biotechnological processes presently impaired from limited tolerances could similarly benefit from this research.