Development of a Hyperosmotic Stress Inducible Gene Expression System by Engineering the MtrA/MtrB-Dependent NCgl1418 Promoter in Corynebacterium glutamicum

• Published in: Frontiers in microbiology Vol. 12; p. 718511
• Main Authors: Huang, Jingwen, Chen, Jiuzhou, Wang, Yu, Shi, Tuo, Ni, Xiaomeng, Pu, Wei, Liu, Jiao, Zhou, Yingyu, Cai, Ningyun, Han, Shuangyan, Zheng, Ping, Sun, Jibin
• Format: Journal Article
• Language: English
• Published: Frontiers Media S.A 21.07.2021
• Subjects: Corynebacterium glutamicum; fluorescence activated cell sorting; hyperosmotic stress; inducible promoter; lysine; Microbiology; MtrA
• ISSN: 1664-302X; 1664-302X
• DOI: 10.3389/fmicb.2021.718511

Corynebacterium glutamicum is an important workhorse for industrial production of diversiform bioproducts. Precise regulation of gene expression is crucial for metabolic balance and enhancing production of target molecules. Auto-inducible promoters, which can be activated without expensive inducers, are ideal regulatory tools for industrial-scale application. However, few auto-inducible promoters have been identified and applied in C. glutamicum . Here, a hyperosmotic stress inducible gene expression system was developed and used for metabolic engineering of C. glutamicum . The promoter of NCgl1418 (P NCgl1418 ) that was activated by the two-component signal transduction system MtrA/MtrB was found to exhibit a high inducibility under hyperosmotic stress conditions. A synthetic promoter library was then constructed by randomizing the flanking and space regions of P NCgl1418 , and mutant promoters exhibiting high strength were isolated via fluorescence activated cell sorting (FACS)-based high-throughput screening. The hyperosmotic stress inducible gene expression system was applied to regulate the expression of lysE encoding a lysine exporter and repress four genes involved in lysine biosynthesis ( gltA , pck , pgi , and hom ) by CRISPR interference, which increased the lysine titer by 64.7% (from 17.0 to 28.0 g/L) in bioreactors. The hyperosmotic stress inducible gene expression system developed here is a simple and effective tool for gene auto-regulation in C. glutamicum and holds promise for metabolic engineering of C. glutamicum to produce valuable chemicals and fuels.