Enhanced production of l-sorbose in an industrial Gluconobacter oxydans strain by identification of a strong promoter based on proteomics analysis

Abstract

Gluconobacter oxydans is capable of rapidly incomplete oxidation of many sugars and alcohols, which means the strain has great potential for industrial purposes. Strong promoters are one of the essential factors that can improve strain performance by overexpression of specific genes. In this study, a pipeline for screening strong promoters by proteomics analysis was established. Based on the procedure, a new strong promoter designated as P B932_2000 was identified in G. oxydans WSH-003. The promoter region was characterized based on known genome sequence information using BPROM. The strength of P B932_2000 was further assessed by analysis of enhanced green fluorescent protein (egfp) expression and comparison with egfp expression by two commonly used strong promoters, P E. coli_tufB and P G. oxydans_tufB . Both quantitative real-time PCR and fluorescence intensities for egfp gene expression showed that P B932_2000 promoter is stronger than the other two. Overexpression of D-sorbitol dehydrogenase (sldh) by P B932_2000 in G. oxydans WSH-003 enhanced the titer and productivity of L-sorbose synthesis from D-sorbitol by 12.0 % and 33.3 %, respectively. These results showed that proteomics analysis is an efficient way to identify strong promoters. The isolated promoter P B932_2000 could further facilitate the metabolic engineering of G. oxydans.

Isolating promoters from Corynebacterium ammoniagenes ATCC 6871 and application in CoA synthesis

“…Studies have shown that strong promoters are usually obtained from the promoters of essential genes whose transcription levels are presumed to be high and constant [17]. In addition, we found that some commonly-used strong promoters such as P dnak in Gluconobacter oxydans and P gro in Corynebacterium glutamicum are promoters of molecular chaperones [18,19]. In this study, the promoters for 20 genes with high transcription levels and 6 molecular chaperone-encoding genes are identified by bioinformatic methods and their activities were further examined experimentally.…”

Enzymes of industrial interest

“…The bioconversion of D-sorbitol to L-sorbose by Gluconobacter oxydans is catalyzed by D-sorbitol dehydrogenase, encoded by the sldh gene. Increased expression of this gene resulted in increased bioconversion activity (Hu et al, 2015). The sorbose concentration was increased to 144 g/L at 16 hours.…”

Design of a high‐efficiency synthetic system for l‐asparaginase production in Bacillus subtilis

“…A large number of promoters have previously been used successfully to improve product yield . By replacing the promoter, the production increased from 12% to five times as much . However, even under the same promoter, gene expression differed due to differences in translation rates .…”

“…By replacing the promoter, the production increased from 12% to five times as much [19,20]. However, even under the same promoter, gene expression differed due to differences in translation rates [21].…”