Genome-wide targets identification by CRISPRi-Omics for high-titer production of free fatty acids in Escherichia coli

Abstract

AbstractTo construct a superior microbial cell factory for chemical synthesis, a major challenge is to fully exploit cell potential via identifying and engineering beneficial gene targets in the sophisticated metabolic networks. Here, we develop an approach that integrates CRISPR interference (CRISPRi) to readily modulate genes expression and omics analyses to identify potential targets in multiple cellular processes, enabling systematical discovery of beneficial chromosomal gene targets that can be engineered to optimize free fatty acids (FFAs) production in Escherichia coli. We identify 56 beneficial genes via synergistic CRISPRi-Omics strategy, including 46 novel targets functioning in cell structure and division, and signaling transduction that efficiently facilitate FFAs production. Upon repressing ihfA and overexpressing aidB and tesA’ in E. coli, the recombinant strain LihfA-OaidB results in a FFAs titer of 21.6 g L-1 in fed-batch fermentation, which, to our best knowledge, is the maximum FFAs titer by the recombinant E. coli reported to date.