Hydrophobic anion activation of human liver chi chi alcohol dehydrogenase.
Class III alcohol dehydrogenase (chi chi-ADH) from human liver binds both ethanol and acetaldehyde so poorly that their Km values cannot be determined, even at ethanol concentrations up to 3 M. However, long-chain carboxylates, e.g., pentanoate, octanoate, deoxycholate, and other anions, substantially enhance the binding of ethanol and other substrates and hence the activity of class III ADH up to 30-fold. Thus, in the presence of 1 mM octanoate, ethanol displays Michaelis-Menten kinetics. The degree of activation depends on the size both of the substrate and of the activator; generally, longer, negatively charged activators result in greater activation. At pH 10, the activator binds to the E-NAD+ form of the enzyme to potentiate substrate binding. Pentanoate activates methylcrotyl alcohol oxidation and methylcrotyl aldehyde reduction 14- and 30-fold, respectively. Such enhancements of both oxidation and reduction are specific for class III ADH; neither class I nor class II shows this effect. The implications as to the nature of the physiological substrate(s) of class III ADH are discussed in light of the recent finding that this ADH and glutathione-dependent formaldehyde dehydrogenase are identical. A new rapid purification procedure for chi chi-ADH is presented.
Moulis
J M
JM
Center for Biochemical and Biophysical Sciences and Medicine, Harvard Medical School, Boston, Massachusetts 02115.
Holmquist
B
B
Vallee
B L
BL
D000445:Aldehyde Oxidoreductases / Q000378:metabolism*; D000447:Aldehydes / Q000378:metabolism; D000838:Anions / Q000378:metabolism; D001665:Binding Sites; D004789:Enzyme Activation; D000431:Ethanol / Q000378:metabolism; D006801:Humans; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D008099:Liver / Q000201:enzymology*; D011487:Protein Conformation; D012995:Solubility; D013379:Substrate Specificity / Q000187:drug effects; D014631:Valerates / Q000494:pharmacology
D000447:Aldehydes; D000838:Anions; D014631:Valerates; D000431:Ethanol; C020145:formaldehyde dehydrogenase (glutathione); D000445:Aldehyde Oxidoreductases