Pleiotropic Global Regulator Lrp of Free-Living Nitrogen-Fixing Ethanologenic Bacterium Couples Carbon–Nitrogen Metabolism

Rational design and construction of robust microbial cell factories (MCFs) require comprehensive information about metabolic and regulatory networks. However, this is hindered by limited information and toolkits for nonmodel industrial microorganisms. Here, we investigated the function of pleiotropic global regulator Lrp in the nitrogen-fixing ethanologenic bacterium Zymomonas mobilis. Although the deletion of lrp significantly reduced cell growth under both feast and famine conditions, the nitrogen fixation efficiency was enhanced. This could be attributed to the upregulation of nitrogen fixation regulators nifA and glnB in Δlrp based on omics analyses and electrophoretic mobility shift assay results. Additionally, Lrp exhibits a broad regulatory role in regulating genes involved in nitrogen and carbon metabolism, including those associated with ammonia assimilation, amino acid metabolism, the TCA cycle, and one-carbon metabolism. Moreover, independent, concerted, and reciprocal regulatory modes of Lrp upon leucine supplementation were proposed. Notably, lrp deletion was also confirmed to effectively enhance nitrogen fixation capability in a d-lactate-producing Z. mobilis strain and the model legume-rhizobia symbiotic bacterium Mesorhizobium loti. This study thus provides new insights into Lrp-mediated regulation of nitrogen fixation and carbon–nitrogen metabolisms as well as strategies to release the tightly controlled nitrogen fixation, which contributes to the design and construction of robust and efficient MCFs.