Characterisation of a recombinant NADP-dependent glycerol dehydrogenase from Gluconobacter oxydans and its application in the production of L-glyceraldehyde.
The acetic acid bacterium Gluconobacter oxydans has a high potential for oxidoreductases with a variety of different catalytic abilities. One putative oxidoreductase gene codes for an enzyme with a high similarity to the NADP+-dependent glycerol dehydrogenase (GlyDH) from Hypocrea jecorina. Due to this homology, the GlyDH (Gox1615) has been cloned, over-expressed in Escherichia coli, purified and characterised. Gox1615 shows an apparent native molecular mass of 39 kDa, which corresponds well to the mass of 37.213 kDa calculated from the primary structure. From HPLC measurements, a monomeric structure can be deduced. Kinetic parameters and the dependence of the activity on temperature and pH were determined. The enzyme shows a broad substrate spectrum in the reduction of different aliphatic, branched and aromatic aldehydes. Additionally, the enzyme has been shown to oxidize a variety of different alcohols. The highest activities were observed for the conversion of D-glyceraldehyde in the reductive and L-arabitol in the oxidative direction. Since high enantioselectivities were observed for the reduction of glyceraldehyde, the kinetic resolution of glyceraldehyde was investigated and found to yield enantiopure L-glyceraldehyde on preparative scale.
Richter
Nina
N
Evocatal GmbH, Merowingerplatz 1 A, 40225 Düsseldorf, Germany.
Neumann
Markus
M
Liese
Andreas
A
Wohlgemuth
Roland
R
Eggert
Thorsten
T
Hummel
Werner
W
D000595:Amino Acid Sequence; D000818:Animals; D020600:Gluconobacter oxydans / Q000201:enzymology*; D005985:Glyceraldehyde / Q000378:metabolism*; D008969:Molecular Sequence Data; D009249:NADP / Q000378:metabolism; D010084:Oxidation-Reduction; D011817:Rabbits; D011994:Recombinant Proteins / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism; D017386:Sequence Homology, Amino Acid; D013379:Substrate Specificity; D013401:Sugar Alcohol Dehydrogenases / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*
D011994:Recombinant Proteins; D005985:Glyceraldehyde; D009249:NADP; D013401:Sugar Alcohol Dehydrogenases; C020673:glycerol 2-dehydrogenase (NADP+)