Harold J. Rhodes

DEUTERIUM ISOTOPE EFFECTS IN ENZYMATIC TRANSAMINATION.

Abstract The ionic fraction from the hydrolysate of residual cell walls of Scenedesmus obliquus grown in a medium containing 99.6% 2H2O was subjected to ion-exchange chromatography for the fractionation of fully deuterated l -glutamic acid. l -[2-2H]-Glutamic acid and l -[3,3,4,4-2H4]glutamic acid were synthesized from l -glutamic acid and l -[2,3,4,4-2H5]glutamic acid, respectively. A kinetic study of enzymatic transamination was conducted using the four l -glutamic acid analogs ( l -glutamic acid and three deuterated analogs) as the substrates. The rate of transamination was followd by photometric measurements at 280 mμ. Maximum velocity determinations, based on the binary mechanism of the enzymatic transamination, were performed on each of the l -glutamic acid analogs. The deuterium isotope effect reflected in the rate of transmission was estimated through maximum velocity analysis. The compound with deuterium in the α-position showed an isotope effect of 1.85. A similar isotope effect was observed for the compound with deuterium at the α-, β- and γ-carbons. The data indicate that the bond to the α-carbon and not to the β-carbon is ruptured during transamination. It is clearly demonstrated, therefore, that an α-elimination mechanism is involved in the enzymatic transamination.