The purification and some properties of the Mg(2+)-activated cytosolic aldehyde dehydrogenase of Saccharomyces cerevisiae.
A purification procedure has been developed for the cytosolic aldehyde dehydrogenase of Saccharomyces cerevisiae that yields homogeneous enzyme. The enzyme seems to be a tetramer of identical 58 kDa subunits. The enzyme reaction is strongly stimulated by Mg2+ at low NADP+ concentrations but there is no absolute requirement for bivalent cations. The kinetics of the reaction have been studied in the presence and absence of MgCl2. NADP+ binding studies of the quenching of protein fluorescence in the presence and absence of MgCl2 show that the effect of Mg2+ is to increase the affinity of the enzyme for NADP+ by approx. 100-fold. NADP+ binding causes a slow conformational change in the enzyme and converts the enzyme from the inactive or low-activity form in which it is isolated into the fully active form. This conformational change seems to explain the marked lag-phases seen in enzyme assays. The enzyme is strongly inhibited by disulfiram and pyridoxal 5-phosphate.
Dickinson
F M
FM
Department of Applied Biology, University of Hull, U.K.
D000079:Acetaldehyde / Q000378:metabolism; D000444:Aldehyde Dehydrogenase / Q000737:chemistry / Q000302:isolation & purification* / Q000378:metabolism*; D001665:Binding Sites; D003600:Cytosol / Q000201:enzymology; D004221:Disulfiram / Q000494:pharmacology; D004789:Enzyme Activation; D004791:Enzyme Inhibitors / Q000494:pharmacology; D007527:Isoenzymes / Q000737:chemistry / Q000302:isolation & purification* / Q000378:metabolism*; D007700:Kinetics; D008274:Magnesium / Q000378:metabolism; D008970:Molecular Weight; D009249:NADP / Q000378:metabolism; D010084:Oxidation-Reduction; D011487:Protein Conformation; D011732:Pyridoxal Phosphate / Q000494:pharmacology; D012441:Saccharomyces cerevisiae / Q000201:enzymology*
D004791:Enzyme Inhibitors; D007527:Isoenzymes; D009249:NADP; D011732:Pyridoxal Phosphate; D000444:Aldehyde Dehydrogenase; D000079:Acetaldehyde; D008274:Magnesium; D004221:Disulfiram