Human alcohol dehydrogenase (ADH) constitutes a complex family. Class IV ADH (ADH4) is characteristic in its epithelial expression in the aerodigestive tract and high V(max) and K(m) for oxidation of ethanol. ADH4 exhibits the highest catalytic efficiency for retinol oxidation in human ADH family. Initial velocity, product inhibition, and dead-end inhibition studies indicate that ADH4, when functioning as ethanol dehydrogenase, conforms to an ordered sequential mechanism with coenzyme binding first and releasing last in catalytic cycle. When functioning as retinol dehydrogenase, the mechanism of ADH4 deduced from steady-state kinetic and equilibrium-binding studies is best described as a rapid equilibrium random mechanism with two dead-end ternary complex for retinol oxidation and a rapid equilibrium ordered mechanism with one dead-end ternary complex for retinal reduction, a unique mechanistic form for zinc-containing ADHs in the medium chain dehydrogenase/reductase superfamily. Kinetic and genetic studies support the proposal that ADH4 may play two important physiological roles, i.e., as a major contributor to first-pass metabolism of ethanol in stomach as well as involvement in the synthesis of retinoic acid, a hormonal ligand controlling a nuclear receptor signaling pathway that regulates growth, development, and epithelial maintenance. Quantitative simulation studies indicate that retinol metabolism through ADH pathway can be inhibited to a significant extent during alcohol consumption. The perturbation of retinoic acid synthesis by ethanol may underlie the pathogenesis of fetal alcohol syndrome and alcohol-related upper digestive tract cancer.