Mechanism of inhibition of aldehyde dehydrogenase by citral, a retinoid antagonist

Low concentrations of citral (3,7-dimethyl-2,6-octadienal), an inhibitor of retinoic acid biosynthesis, inhibited El, E2 and E3 isozymes of human aldehyde dehydrogenase (EC 1.2.1.3). The inhibition was reversible on dilution and upon long incubation in the presence of NAD(+); it occurred with simultaneous formation of NADH and of geranic acid. Thus, citral is an inhibitor and also a substrate. K-m, values for citral were 4 mu M for E1, 1 mu M for E2 and 0.1 mu M for E3; V-max values were highest for E1 (73 nmol.min(-1).mg(-1)), intermediate for E2, (17 nmol.min(-1).mg(-1)) and lowest (0.07 nmol.min(-1).mg(-1)) for the E3 isozyme. Citral is a 1 : 2 mixture of isomers: cis isomer neral and trans isomer, geranial; the latter structurally resembles physiologically important retinoids. Both were utilized by all three isozymes; a preference for the trans isomer, geranial, was observed by HPLC and by enzyme kinetics. With the El isozyme, both geranial and neral, and with the E2 isozyme, only neral obeyed Michaelis-Menten kinetics. With the E2 isozyme and geranial sigmoidal saturation curves were observed with S-0.5 of approximate to 50 nM; the n-values of 2-2.5 indicated positive cooperativity. Geranial was a better substrate and a better inhibitor than neral. The low V-max, which appeared to be controlled by either the slow formation, or decomposition via the hydride transfer, of the thiohemiacetal reaction intermediate, makes citral an excellent inhibitor whose selectivity is enhanced by low K-m values. The V-max for citral with the E1 isozyme was higher than those of the E2 and E3 isozymes which explains its fast recovery following inhibition by citral and suggests that El map be the enzyme involved in vivo citral metabolism.