A pH-controlled fed-batch process can overcome inhibition by formate in NADH-dependent enzymatic reductions using formate dehydrogenase-catalyzed coenzyme regeneration.
The NAD-dependent, formate dehydrogenase-catalyzed oxidation of formate anion into CO2 is known as the method for the regeneration of NADH in reductive enzymatic syntheses. Inhibition by formate and inactivation by alkaline pH-shift that occurs when oxidation of formate is carried out at pH approximately 7.0 may, however, hamper the efficient application of this NADH recycling reaction. Here, we have devised a fed-batch process using pH-controlled feeding of formic acid that can overcome enzyme inhibition and inactivation. The reaction pH is thus kept constant by addition of acid, and formate dehydrogenase is supplied continuously with substrate as required, but the concentration of formate is maintained at a constant, non- or weakly inhibitory level throughout the enzymatic conversion, thus enabling a particular NADH-dependent dehydrogenase to operate stably and at high reaction rates. For xylitol production from xylose using yeast xylose reductase (Ki,Formate 182 mM), a fed-batch conversion of 0.5M xylose yielded productivities of 2.8 g (L h)-1 that are three-fold improved when contrasted to a conventional batch reaction that employed equal initial concentrations of xylose and formate.
Neuhauser
W
W
Division of Biochemical Engineering, Universität für Bodenkultur Wien (BOKU), Muthgasse 18, A-1190 Wien, Austria.
Steininger
M
M
Haltrich
D
D
Kulbe
K D
KD
Nidetzky
B
B
D000449:Aldehyde Reductase / Q000378:metabolism*; D002175:Candida / Q000201:enzymology; D005560:Formate Dehydrogenases / Q000037:antagonists & inhibitors / Q000378:metabolism*; D005561:Formates / Q000494:pharmacology*; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D008956:Models, Chemical; D009243:NAD / Q000378:metabolism*; D010084:Oxidation-Reduction; D014993:Xylitol / Q000096:biosynthesis*; D014994:Xylose / Q000378:metabolism
D005561:Formates; D009243:NAD; C030544:formic acid; D014994:Xylose; D000449:Aldehyde Reductase; D005560:Formate Dehydrogenases; D014993:Xylitol