The inhibition of brain aryl acylamidase by 5-hydroxytryptamine and acetylcholine

Abstract

Sheep brain aryl acylamidase (aryl-acylamide amidohydrolase. EC 3.5.1.13) was partially purified. Of a number of amines tested at 1 mM the enzyme was maximally inhibited by 5-hydroxytryp-tamine (5-HT) and to a lesser extent by tryptamine and practically unaffected by tyramine, histamine, noradrenaline, dopamine and benzylamine and a number of amino acids including tryptophan. Choline derivatives at 1 mM were inhibitory to the enzyme in the order of butyrylcholine > succinylcholine > benzoylcholine > choline > acetylcholine > acetylthiocholine > acetyl-β-methylcholine > pro-pionylcholine. The inhibition by 5-HT and acetylcholine was further studied. Both inhibit the enzyme in a noncompetitive manner. The inhibition could be reversed by removal of the inhibitors from the enzyme by gel filteration. A number of metal ions, EDTA, high concentrations of sodium chloride. the thiol reagents p-chloromercuribenzoate, N-ethyl-maleimide and iodoacetamide were found to have no effect on the inhibition. Dithiothereitol as well as neuraminidase treatment did not alter the extent of inhibition of the enzyme by 5-HT or acetylcholine. Nitration of the enzyme with tetranitromethane led to approximately a 50 per cent drop in enzyme activity as well as a significant decrease in the extent of inhibition by 5-HT and acetylcholine. This suggested the possibility of the involvement of tyrosine residues both at the catalytic site as well as at the site(s) of inhibition by 5-HT and acetylcholine. Mixed inhibitor studies favoured a common inhibition site for both 5-HT and acetylcholine on the brain enzyme. The sheep liver enzyme was not inhibited by either 5-HT or acetylcholine.