Deuterium isotope effects on the enzymatic oxidative deamination of trace amines

Abstract

The steady-state kinetics of the oxidative deamination of some trace amines [ p-tyramine ( p-TA), m-tyramine ( m-TA) and β-phenylethylamine (PE)] and the same trace amines containing deuterium in their side-chain (i.e. αα-d 2 and ββ-d 2) have been assessed using rat liver mitochondrial monoamine oxidase (MAO) incubated at 37° in 0.05 M phosphate buffer (pH 7.5). In these in vitro reactions, a considerable reduction in deamination occurred when the deuterium substitution was in the α position. In addition, the isotope effect was found to be related to hydroxyl substitution on the phenyl ring. The apparent ( V/ K) H /( V/ K) D ratios were 4.44, 4.24 and 2.06 for p-TA, m-TA and PE respectively. We have confirmed that the cleavage of the C—H bond at the α position is involved in the rate-limiting step of the enzymatic deamination. In the case where the deuterium substitution was in the ββ position, a slight enhancement of deamination occurred with the ( V/ K) H /( V/ K) D ratio becoming 0.89, 0.86 and 0.95, respectively, for p-TA, m-TA and PE. The selective inhibition of the deamination of the αα-deuterated amines by the specific MAO inhibitors clorgyline (type A) and deprenyl (type B) was not different from that of the corresponding non-deuterated trace amines. The isotope effect was found to be somewhat greater at lower temperatures. Using mixed substrates (i.e. trace amine + corresponding deutrated trace amine), the deuterated amines were observed to exhibit a weak inhibitory effect due simply to competition for the same site on the enzyme.