The association of the serotonin-sensitive aryl acylamidase with acetylcholinesterase in the monkey brain

Abstract

The serotonin-sensitive aryl acylamidase was partially purified from monkey brain. The aryl acylamidase activity was inhibited by serotonin (K_i = 0.425 mM) and tryptamine (K_i = 3.6 mM) but not by a number of other amines. It was also inhibited by acetylcholine (K_i = 2 mM) and its analogues and homologues. The relationship of aryl acylamidase to acetylcholinesterase was examined. The ratios of specific activities of aryl acylamidase and acetylcholinesterase in the different steps of purification were approximately constant and the percentage recoveries of both enzyme activities were comparable. Elution profiles of both enzyme activities from concanavalin-A-Sepharose, Sephadex G-200 and DEAE-Sephadex A-25 columns were similar. Both enzyme activities migrated in a similar fashion on gel electrophoresis in different percentage gels for different time intervals. Both enzymes showed similar distribution in the various anatomical regions and in the different subcellular fractions of monkey brain. Eserine and neostigmine, potent competitive inhibitors of acetylcholinesterase also potently inhibited aryl acylamidase in a non-competitive manner. Inhibition of both enzymes was 100% at 10 μM of both the inhibitors. Tetraisopropylpyrophosphoramide, a selective inhibitor of pseudo-cholinesterase, did not inhibit either the brain acetylcholinesterase or aryl acylamidase at 10μM. Serotonin inhibited acetylcholinesterase only at concentrations above 10 mM. Dixon plots of one inhibitor in the presence of additional inhibitors indicated that serotonin, butyrylcholine, acetylcholine, neostigmine and eserine (all non-competitive inhibitors of aryl acylamidase) acted at the same site as inhibitors of aryl acylamidase. The serotonin-insensitive aryl acylamidase of monkey liver was also insensitive to acetylcholine and eserine and was not found associated with acetylcholinesterase [A. Oommen and A. S. Balasubramanian (1978) Biochem. Pharmacol. 27, 891-895]. Erythrocyte membrane known to contain true cholinesterase had aryl acylamidase activity sensitive to serotonin, acetylcholine and eserine. All these considerations suggest that the serotonin-sensitive aryl acylamidase activity is a property of true cholinesterase. Although some of the experimental results would suggest that in the monkey brain the two activities may be associated with the same protein with two different active sites further experiments are needed to confirm this.