Construction Of Escherichia Coli Cell Factory For Lycopene Production And Its Fermentation Optimization

Lycopene belongs to carotenoids family, which has lots of applications in pharmaceuticals, neutraceuticals, cosmetics and food industries. Lycopene can prevent oxidation of low density lipoprotein and cholesterol, thereby playing an important role in human heath as biological antioxidants. In recent years, the interest in producing lycopene by microbial fermentation with recombinant E. coli has increased.Firstly, a previously constructed β-carotene-producing strain CAR001was engineered for lycopene production by deleting zeaxanthin glucosyltransferase (crtX) and lycopene β-cyclase (crtY) genes in the crtEXYIB gene operon through homologous recombination. The resulting strain LYC001produced10.49mg/L lycopene with a yield of6.52mg/g DCW.Secondly, the a-ketoglutarate dehydrogenase (sucAB), succinate dehydrogenase (sdhABCD) and transaldolase B (talB) genes within central metabolic modules were engineered to increase ATP and NADPH supplies. Modulation of sucAB gene of TCA module caused up to64%increase of lycopene yield, while modulation of sucAB and talB genes caused up to70%increase. Modulating these three key genes led to76%increase of lycopene yield. The resulting strain LYC005produced18.91mg/L lycopene with a yield of11.53mg/g DCW. It was suggested that engineering TCA and PPP modules could significantly improve lycopene production.Thirdly, lycopene production was improved by increasing metabolic flux towards MEP pathway through RBS library modulation of key isoprenoid genes to increase isopentenyl diphosphate (IPP) and dimethylallyl diphosphate (DMAPP) supply. In this work, modulating1-deoxy-D-xylulose-5-phosphate synthase (dxs), isopentenyl diphosphate isomerase (idi) and crt genes with RBS libraries led to13%,13%and14%improvement of lycopene yield compared to parent strain, respectively. Combined modulation of these three genes with RBS libraries resulted in32%improvement of lycopene yield. The resulting strain LYC010produced26.33mg/L lycopene with a yield of15.22mg/g DCW. Our results demonstrated that modulating gene expression with RBS librariy was better than using several regulatory parts with fixed strengths.Through combined engineering of MEP, lycopene synthesis and central metabolic modules, a genetically stable E. coli strain LYC010was obtained. Fed-batch fermentation of this strain was performed, which produced3.52g/L lycopene with a yield of50.57mg/g DCW.