Re-face stereospecificity at C4 of NAD(P) for alcohol dehydrogenase from Methanogenium organophilum and for (R)-2-hydroxyglutarate dehydrogenase from Acidaminococcus fermentans as determined by H-1-NMR spectroscopy

The two diastereotopic protons at C4 of NAD(P)H are seen separately in H-1-NMR spectra. This fact was used to determine the stereospecificity at C4 of NAD(P) for the NADP-dependent alcohol dehydrogenase from Methanogenium organophilum and for the NAD-dependent (R)-2-hydroxyglutarate dehydrogenase from Acidaminococcus fermentans. The reduction of NADP(+) with [H-2(6)]ethanol was found to yield (4R)-[4-H-2(1)]NADPH and the oxidation of (4R)-[4-H-2(1)]NADH with 2-oxoglutarate to yield unlabelled [4-H-1]NAD(+). These results indicate that both enzymes are Re-face stereospecific at C4 of the pyridine nucleotides.