Kang Aram, Chang Matthew Wook
Division of Chemical and Biomolecular Engineering, School of Chemical and Biomedical Engineering, Nanyang Technological University, 637459, Singapore.
Mol Biosyst. 2012 Apr;8(4):1350-8. doi: 10.1039/c2mb05441h. Epub 2012 Feb 10.
Microbial tolerance to hydrocarbons has been studied in an effort to improve the productivity of biochemical processes and to enhance the efficiency of hydrocarbon bioremediation. Despite these studies, few attempts have been made to design rational strategies to improve microbial tolerance to hydrocarbons. Herein, we present an engineering framework that enables us to harness our understanding of genetic regulatory networks to improve hydrocarbon tolerance. In this study, isooctane was used as a representative hydrocarbon due to its use in petroleum refining and in biochemical processes. To increase isooctane tolerance, we first identified essential transcriptional determinants and genetic regulatory networks underlying cellular responses to isooctane in Escherichia coli using genome-wide microarray analysis. Based on functional transcriptome and bioinformatics analysis, a range of combinations of transcription factors whose activity was predictably perturbed by isooctane were knocked out and overexpressed to reconstitute the regulatory networks. We demonstrated that the reconstitution of the regulatory networks led to a significant improvement in isooctane tolerance, and especially, engineered E. coli strains lacking and overexpressing some of the perturbed transcription factors showed 3- to 5-fold improvement. This microbe with high tolerance to isooctane can be harnessed for biochemical processes, fuel oil bioremediation and metabolic engineering for biofuel production. Furthermore, we envision that the engineering framework employed to improve the tolerance in this study can be exploited for developing other microbes with desired phenotypes.
为了提高生化过程的生产率并增强烃类生物修复的效率，人们对微生物对烃类的耐受性进行了研究。尽管有这些研究，但很少有人尝试设计合理的策略来提高微生物对烃类的耐受性。在此，我们提出了一个工程框架，使我们能够利用对基因调控网络的理解来提高对烃类的耐受性。在本研究中，异辛烷因其在石油精炼和生化过程中的应用而被用作代表性烃类。为了提高对异辛烷的耐受性，我们首先使用全基因组微阵列分析确定了大肠杆菌中对异辛烷细胞反应的基本转录决定因素和基因调控网络。基于功能转录组和生物信息学分析，敲除并过表达了一系列其活性可被异辛烷预测性干扰的转录因子组合，以重建调控网络。我们证明，调控网络的重建导致异辛烷耐受性显著提高，特别是，缺失和过表达一些受干扰转录因子的工程大肠杆菌菌株显示出3至5倍的改善。这种对异辛烷具有高耐受性的微生物可用于生化过程、燃料油生物修复和生物燃料生产的代谢工程。此外，我们设想，本研究中用于提高耐受性的工程框架可用于开发具有所需表型的其他微生物。
