Fig. 6: A Cra-regulated pBAD promoter improves overproduction of carotenoids.

a Metabolic map of the E. coli methylerythritol phosphate (MEP) pathway and the synthetic carotenoid pathway. Expression of the first enzymes in the MEP pathway (1-deoxy-d-xylulose-5-phosphate synthase, Dxs; and 1-deoxy-d-xylulose 5-phosphate reductoisomerase, Dxr) was controlled with a pBAD promoter (plasmid pController) or with a pBAD-Cra promoter (plasmid pController-Cra). The carotenoid pathway consists of five enzymes from P. ananatis: geranylgeranyl diphosphate synthase (CrtE), phytoene synthase (CrtB), phytoene dehydrogenase (CrtI), lycopene beta-cyclase (CrtY), and beta-carotene hydroxylase (CrtZ). These five genes were expressed under the control of a native constitutive promoter from P. ananatis and one additional dxs gene from E. coli (plasmid pCarotenoid). b Plasmids pCarotenoid and pController were co-expressed in E. coli (pBAD carotenoid strain). Both strains were grown in a plate reader at different induction levels for Dxs and Dxr (0, 0.1, 0.3, 0.5, 1, and 2% ara). The total carotenoid content was measured after 24 h. Growth curves and dots show means of n = 2 cultures, and gray shaded areas show the difference between the two cultures. GAP glyceraldehyde-3-phosphate, DXP 1-deoxy-d-xylulose-5-phosphate, MEP 2-C-methyl-d-erythritol-4-phosphate, DMAPP dimethylallyl diphosphate, IPP isopentenyl diphosphate, GPP geranyl diphosphate, FPP farnesyl diphosphate, GGPP geranylgeranyl diphosphate. Source data are provided in the Source Data file.