Expression of glf Z.m. increases D-mannitol formation in whole cell biotransformation with resting cells of Corynebacterium glutamicum.
A recombinant oxidation/reduction cycle for the conversion of D-fructose to D-mannitol was established in resting cells of Corynebacterium glutamicum. Whole cells were used as biocatalysts, supplied with 250 mM sodium formate and 500 mM D-fructose at pH 6.5. The mannitol dehydrogenase gene (mdh) from Leuconostoc pseudomesenteroides was overexpressed in strain C. glutamicum ATCC 13032. To ensure sufficient cofactor [nicotinamide adenine dinucleotide (reduced form, NADH)] supply, the fdh gene encoding formate dehydrogenase from Mycobacterium vaccae N10 was coexpressed. The recombinant C. glutamicum cells produced D-mannitol at a constant production rate of 0.22 g (g cdw)(-1) h(-1). Expression of the glucose/fructose facilitator gene glf from Zymomonas mobilis in C. glutamicum led to a 5.5-fold increased productivity of 1.25 g (g cdw)(-1) h(-1), yielding 87 g l(-1) D-mannitol from 93.7 g l(-1) D-fructose. Determination of intracellular NAD(H) concentration during biotransformation showed a constant NAD(H) pool size and a NADH/NAD(+) ratio of approximately 1. In repetitive fed-batch biotransformation, 285 g l(-1) D-mannitol over a time period of 96 h with an average productivity of 1.0 g (g cdw)(-1) h(-1) was formed. These results show that C. glutamicum is a favorable biocatalyst for long-term biotransformation with resting cells.
Bäumchen
Carsten
C
Institut für Biotechnologie 1, Forschungszentrum Jülich GmbH, 52425, Jülich, Germany. st.bringer-meyer@fz-juelich.de
Bringer-Meyer
Stephanie
S
D001426:Bacterial Proteins / Q000235:genetics* / Q000378:metabolism*; D001483:Base Sequence; D001709:Biotechnology; D001711:Biotransformation; D048230:Corynebacterium glutamicum / Q000235:genetics* / Q000378:metabolism*; D017931:DNA Primers / Q000235:genetics; D004269:DNA, Bacterial / Q000235:genetics; D005560:Formate Dehydrogenases / Q000235:genetics / Q000378:metabolism; D005632:Fructose / Q000378:metabolism; D015870:Gene Expression; D005798:Genes, Bacterial; D007934:Leuconostoc / Q000201:enzymology / Q000235:genetics; D008353:Mannitol / Q000378:metabolism*; D008354:Mannitol Dehydrogenases / Q000235:genetics / Q000378:metabolism; D009154:Mutation; D009161:Mycobacterium / Q000235:genetics / Q000378:metabolism; D009243:NAD / Q000378:metabolism; D010084:Oxidation-Reduction; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D017355:Zymomonas / Q000235:genetics / Q000378:metabolism
D001426:Bacterial Proteins; D017931:DNA Primers; D004269:DNA, Bacterial; D011994:Recombinant Proteins; C086512:glucose facilitator protein, Zymomonas; D009243:NAD; D005632:Fructose; D008353:Mannitol; D008354:Mannitol Dehydrogenases; D005560:Formate Dehydrogenases