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- Title
Enhanced production of GDP- l-fucose by overexpression of NADPH regenerator in recombinant Escherichia coli.
- Authors
Won-Heong Lee; Young-Wook Chin; Nam Soo Han; Myoung-Dong Kim; Jin-Ho Seo
- Abstract
Biosynthesis of guanosine 5′-diphosphate- l-fucose (GDP- l-fucose) requires NADPH as a reducing cofactor. In this study, endogenous NADPH regenerating enzymes such as glucose-6-phosphate dehydrogenase (G6PDH), isocitrate dehydrogenase (Icd), and NADP-dependent malate dehydrogenase (MaeB) were overexpressed to increase GDP- l-fucose production in recombinant Escherichia coli. The effects of overexpression of each NADPH regenerating enzyme on GDP- l-fucose production were investigated in a series of batch and fed-batch fermentations. Batch fermentations showed that overexpression of G6PDH was the most effective for GDP- l-fucose production. However, GDP- l-fucose production was not enhanced by overexpression of G6PDH in the glucose-limited fed-batch fermentation. Hence, a glucose feeding strategy was optimized to enhance GDP- l-fucose production. Fed-batch fermentation with a pH-stat feeding mode for sufficient supply of glucose significantly enhanced GDP- l-fucose production compared with glucose-limited fed-batch fermentation. A maximum GDP- l-fucose concentration of 235.2 ± 3.3 mg l, corresponding to a 21% enhancement in the GDP- l-fucose production compared with the control strain overexpressing GDP- l-fucose biosynthetic enzymes only, was achieved in the pH-stat fed-batch fermentation of the recombinant E. coli overexpressing G6PDH. It was concluded that sufficient glucose supply and efficient NADPH regeneration are crucial for NADPH-dependent GDP- l-fucose production in recombinant E. coli.
- Subjects
BIOSYNTHESIS; ESCHERICHIA coli; GUANOSINE triphosphatase; GLUCOSE; REGENERATION (Biology); FERMENTATION
- Publication
Applied Microbiology & Biotechnology, 2011, Vol 91, Issue 4, p967
- ISSN
0175-7598
- Publication type
Article
- DOI
10.1007/s00253-011-3271-x