Effects of NADH kinase on NADPH-dependent biotransformation processes in Escherichia coli.
Sufficient supply of NADPH is one of the most important factors affecting the productivity of biotransformation processes. In this study, construction of an efficient NADPH-regenerating system was attempted using direct phosphorylation of NADH by NADH kinase (Pos5p) from Saccharomyces cerevisiae for producing guanosine diphosphate (GDP)-L-fucose and ε-caprolactone in recombinant Escherichia coli. Expression of Pos5p in a fed-batch culture of recombinant E. coli producing GDP-L-fucose resulted in a maximum GDP-L-fucose concentration of 291.5 mg/l, which corresponded to a 51 % enhancement compared with the control strain. In a fed-batch Baeyer-Villiger (BV) oxidation of cyclohexanone using recombinant E. coli expressing Pos5p, a maximum ε-caprolactone concentration of 21.6 g/l was obtained, which corresponded to a 96 % enhancement compared with the control strain. Such an increase might be due to the enhanced availability of NADPH in recombinant E. coli expressing Pos5p. These results suggested that efficient regeneration of NADPH was possible by functional expression of Pos5p in recombinant E. coli, which can be applied to other NADPH-dependent biotransformation processes in E. coli.
Department of Agricultural Biotechnology and Center for Agricultural Biomaterials, Seoul National University, Seoul, 151-921, Korea.
Won-Heong
WH
Lee
Jin-Woo
JW
Kim
Eun-Hee
EH
Park
Nam Soo
NS
Han
Myoung-Dong
MD
Kim
Jin-Ho
JH
Seo
D001711:Biotransformation; D003512:Cyclohexanones / Q000378:metabolism; D004926:Escherichia coli / Q000235:genetics* / Q000378:metabolism*; D005818:Genetic Engineering; D024101:Mitochondrial Proteins / Q000235:genetics / Q000378:metabolism*; D009249:NADP / Q000378:metabolism*; D010084:Oxidation-Reduction; D017853:Phosphotransferases (Alcohol Group Acceptor) / Q000235:genetics / Q000378:metabolism*; D029701:Saccharomyces cerevisiae Proteins / Q000235:genetics / Q000378:metabolism*
D003512:Cyclohexanones; D024101:Mitochondrial Proteins; D029701:Saccharomyces cerevisiae Proteins; D009249:NADP; C036468:cyclohexanone; D017853:Phosphotransferases (Alcohol Group Acceptor); C060670:POS5 protein, S cerevisiae