We found a match
Your institution may have rights to this item. Sign in to continue.
- Title
Reductive whole-cell biotransformation with Corynebacterium glutamicum: improvement of NADPH generation from glucose by a cyclized pentose phosphate pathway using pfkA and gapA deletion mutants.
- Authors
Siedler, Solvej; Lindner, Steffen; Bringer, Stephanie; Wendisch, Volker; Bott, Michael
- Abstract
In this study, the potential of Corynebacterium glutamicum for reductive whole-cell biotransformation is shown. The NADPH-dependent reduction of the prochiral methyl acetoacetate (MAA) to the chiral ( R)-methyl 3-hydroxybutyrate (MHB) by an alcohol dehydrogenase from Lactobacillus brevis ( Lbadh) was used as model reaction and glucose served as substrate for the regeneration of NADPH. Since NADPH is mainly formed in the oxidative branch of the pentose phosphate pathway (PPP), C. glutamicum was engineered to redirect carbon flux towards the PPP. Mutants lacking the genes for 6-phosphofructokinase ( pfkA) or glyceraldehyde 3-phosphate dehydrogenase ( gapA) were constructed and analyzed with respect to growth, enzyme activities, and biotransformation performance. Both mutants showed strong growth defects in glucose minimal medium. For biotransformation of MAA to MHB using glucose as reductant, strains were transformed with an Lbadh expression plasmid. The wild type showed a specific MHB production rate of 3.1 mmol h g and a yield of 2.7 mol mol. The ∆ pfkA mutant showed a similar MHB production rate, but reached a yield of 4.8 mol mol, approaching the maximal value of 6 mol mol expected for a partially cyclized PPP. The specific biotransformation rate of the Δ gapA mutant was decreased by 62 % compared to the other strains, but the yield was increased to 7.9 mol mol, which to our knowledge is the highest one reported so far for this mode of NADPH regeneration. As one fourth of the glucose was converted to glycerol, the experimental yield was close to the theoretically maximal yield of 9 mol mol.
- Subjects
BIOTRANSFORMATION (Metabolism); CORYNEBACTERIUM glutamicum; NICOTINAMIDE adenine dinucleotide phosphate; GLUCOSE; PENTOSE phosphate pathway; DELETION mutation; PHOSPHOFRUCTOKINASES; 3-Hydroxybutyric acid
- Publication
Applied Microbiology & Biotechnology, 2013, Vol 97, Issue 1, p143
- ISSN
0175-7598
- Publication type
Article
- DOI
10.1007/s00253-012-4314-7