Engineering Escherichia coli for Carotenoid Biosynthesis

Abstract

This research integrated the isopentenol utilization pathway (IUP) into the E. coli chromosome and utilized various promoters to optimize lycopene production. Additionally, the research evaluated the effects of overexpressing monoglycosyldiacylglycerol synthase (MGS) and diglucosyldiacylglycerol synthase (DGS) from Acholeplasma laidlawii, to induce the formation of intracellular membrane vesicles, potentially increasing the cell’s capacity to store hydrophobic compounds like carotenoids. In a second strategy, the application of knock-out mutants for Braun's lipoprotein (lpp) in E. coli led to the production of extracellular membrane vesicles to avoid intracellular enzyme accumulations. Results demonstrated that integrating the IUP in the chromosome significantly improved lycopene yields compared to traditional pathways and the plasmid system. The overexpression of MGS and DGS resulted in increased intracellular lipid content but did not significantly enhance carotenoid production beyond IUP-expressing strains. Notably, knocking out lpp yielded a 3.3-fold increase in extracellular lycopene production. Overexpressing MEP pathway enzymes, including IDI, GGPPS, CrtI, CrtB, DXS, IspD, IspF, IspG, and IspH in the leaky strain, further boosted lycopene yields. This research underscores the potential of genetic and metabolic engineering to optimize isoprenoid production in microbial hosts, paving the way for more efficient and scalable production methods for these valuable compounds. The findings highlight the efficacy of the IUP and extracellular production strategies in overcoming traditional pathway limitations and enhancing yields, thereby contributing to the broader application of microbial biosynthesis in industrial and pharmaceutical contexts.