[Affinity chromatography of NAD(P)+ dehydrogenases dependent on the concentration of NAD(P)+ bound to ketones or aldehydes].
The use of general ligands such as NAD(P)+ for affinity chromatography of dehydrogenase requires elution with pulses of oxidized or reduced cofactor at suitable concentrations. This method of elution can involve considerable effort before ideal eluent conditions are evolved. We report on a new method of immobilized NAD(P)+ modifications which increases the selectivity of the phase and may be of rather wide application. The immobilized NAD(P)+ is modified by addition of a ketone or an aldehyde substrate of the dehydrogenase which must be purified and becomes a very specific ligand of that dehydrogenase. The adduct of immobilized NAD+ and sodium pyruvate absorbs specifically the lactate dehydrogenase, whereas the adduct of immobilized NAD+ and cyclohexanone adsorbs specifically the horse liver alcohol dehydrogenase. Absorbed dehydrogenases are eluted with NaCl gradients.
D000447:Aldehydes / Q000378:metabolism; D000818:Animals; D002846:Chromatography, Affinity / Q000379:methods; D006736:Horses; D007659:Ketones / Q000378:metabolism; D008099:Liver / Q000201:enzymology; D009247:NADH, NADPH Oxidoreductases / Q000302:isolation & purification*; D009249:NADP / Q000031:analogs & derivatives*; D009252:NADPH Dehydrogenase / Q000302:isolation & purification*
D000447:Aldehydes; D007659:Ketones; D009249:NADP; D009247:NADH, NADPH Oxidoreductases; D009252:NADPH Dehydrogenase
Utilisation pour la chromatographie d'affinité des déshydrogénases NAD(P)+ dépendantes de composes d'addition du NAD(P)+ immobilisé avec les cétones ou les aldéhydes.