Engineering the lycopene synthetic pathway in E. coli by comparison of the carotenoid genes of Pantoea agglomerans and Pantoea ananatis

• Published in: Applied microbiology and biotechnology 2007-02, Vol.74 (1), p.131-139
• Main Authors: Yoon, Sang-Hwal, Kim, Ju-Eun, Lee, Sook-Hee, Park, Hye-Min, Choi, Myung-Suk, Kim, Jae-Yean, Lee, Si-Hyoung, Shin, Yong-Chul, Keasling, Jay D, Kim, Seon-Won
• Format: Article
• Language: English
• Subjects: Archaeoglobus fulgidus; Bacteria; Bacterial Proteins - genetics; Bacterial Proteins - metabolism; Biological and medical sciences; Biotechnology; Biotechnology - methods; Carotenoids; Carotenoids - biosynthesis; Carotenoids - genetics; Carotenoids - metabolism; Cell growth; Culture Media; E coli; Escherichia coli; Escherichia coli - enzymology; Escherichia coli - genetics; Escherichia coli - growth & development; Farnesyltranstransferase - genetics; Farnesyltranstransferase - metabolism; Fundamental and applied biological sciences. Psychology; Gene Expression Regulation, Bacterial; Genetic Engineering - methods; Mevalonic Acid - metabolism; Microbiology; Pantoea - enzymology; Pantoea - genetics; Pantoea agglomerans
• ISSN: 0175-7598; 1432-0614
• DOI: 10.1007/s00253-006-0623-z

The lycopene synthetic pathway was engineered in Escherichia coli using the carotenoid genes (crtE, crtB, and crtI) of Pantoea agglomerans and Pantoea ananatis. E. coli harboring the P. agglomerans crt genes produced 27 mg/l of lycopene in 2YT medium without isopropyl-beta-d-thiogalactopyranoside (IPTG) induction, which was twofold higher than that produced by E. coli harboring the P. ananatis crt genes (12 mg/l lycopene) with 0.1 mM IPTG induction. The crt genes of P. agglomerans proved better for lycopene production in E. coli than those of P. ananatis. The crt genes of the two bacteria were also compared in E. coli harboring the mevalonate bottom pathway, which was capable of providing sufficient carotenoid building blocks, isopentenyl diphosphate (IPP) and dimethylallyl diphosphate (DMAPP), with exogenous mevalonate supplementation. Lycopene production significantly increased using the mevalonate bottom pathway and 60 mg/l of lycopene was obtained with the P. agglomerans crt genes, which was higher than that obtained with the P. ananatis crt genes (35 mg/l lycopene). When crtE among the P. ananatis crt genes was replaced with P. agglomerans crtE or Archaeoglobus fulgidus gps, both lycopene production and cell growth were similar to that obtained with P. agglomerans crt genes. The crtE gene was responsible for the observed difference in lycopene production and cell growth between E. coli harboring the crt genes of P. agglomerans and P. ananatis. As there was no significant difference in lycopene production between E. coli harboring P. agglomerans crtE and A. fulgidus gps, farnesyl diphosphate (FPP) synthesis was not rate-limiting in E. coli.