Thus, after de-bottlenecking the CrtE reaction overexpression of crtB and crtI is beneficial for lycopene overproduction. The maximal lycopene accumulation was 80 fold higher than that of the empty vector control. Lycopene production

was associated with less biomass formation and slowed glucose consumption. In this regard the strain with the highest lycopene production, C. glutamicum ΔcrtEb(pVWEx1-crtE/pEKEx3-crtBI2), stood out. The cells reached the stationary phase after 32 h, exhausted glucose not before 54 h after inoculation and grew only to about half of the biomass concentration (3.7 ± 0.5 mg/ml CDW) as compared to the empty vector control (7.0 ± 0.2 mg/ml CDW). Discussion The GSK2126458 cost synthesis of C50 carotenoids occurs in a restricted number of bacterial species. INK 128 cell line Decaprenoxanthin is the most abundant one and it is the predominant carotenoid of the yellow C. glutamicum. The gene deletion and complementation analysis along with the pathway reconstruction in the multiple deletion strain C. glutamicum ΔΔ corroborates the previous elucidation of decaprenoxanthin biosynthesis in C. glutamicum based on transposon mutants of the strain MJ233C [16] and on

heterologous expression of genes of the crtE-cg0722-crtBIY e Y f Eb cluster in the non-carotenogenic host Escherichia. coli[17]. Furthermore, we have analyzed a hitherto uncharacterized putative second carotenogenic gene cluster of C. glutamicum, crtB2/crtI2-1/crtI2-2, regarding the C50 carotenoid production. For the second

phytoene synthase-like gene, crtB2 (cg2672), annotated in the C. glutamicum genome [25] and postulated to be involved in the squalene synthesis [2], we provide evidence that crtB2 indeed codes for a functional phytoene synthase. Hence, C. glutamicum possesses two functional phytoene synthases, CrtB and CrtB2. The two other open reading frames in the small crt-cluster are annotated as N- and from C-terminal units of a second phytoene desaturase, but experimental confirmation of a phytoene desaturase function could not be obtained. Within the genus Corynebacterium C. glutamicum ATCC 13032 is the only species that possesses a second set of crt genes. The GC content of 54 to 58% of the second crt cluster is similar to the overall GC content of the genome, whereas that of the larger cluster is slightly lower. The genes of the two phytoene synthase paralogs only share 51% identity on the nucleotide level and mobile genetic elements such as IS-elements could not be detected in the vicinity of the two clusters arguing against recent duplication or horizontal gene transfer events. All genome-sequenced corynebacterial species possess a crtI ortholog and most (except C. variabile) also possess a crtB ortholog, either clustered with crtI or elsewhere in the genome. The phylogeny of the crtI gene product reflects the phylogeny of the species. Only the highly related species C. glutamicum and C.