Enhanced production of l-sorbose in an industrial Gluconobacter oxydans strain by identification of a strong promoter based on proteomics analysis
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.
Keywordsl-ascorbic acid Acetic acid bacteria Constitutive promoter Metabolic engineering d-sorbitol
This work was supported by grants from the National High Technology Research and Development Program of China (863 Program, 2012AA022103), the Major State Basic Research Development Program of China (973 Program, 2014CB745103), the Major Program of National Natural Science Foundation of China (21390204), the Program for New Century Excellent Talents in University (NCET-12-0876), the Foundation for the Author of National Excellent Doctoral Dissertation of PR China (FANEDD, 201256), the Priority Academic Program Development of Jiangsu Higher Education Institutions, and the 111 Project (111-2-06).
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