Acetylcholinesterase(AChE)-Catalyzed Hydrolysis of Long-Chain Thiocholine Esters: Shift to a New Chemical Mechanism

Bioanalysis and Biotransformation Research Center, Korea Institute of Science and Technology, Seoul 136-791, KoreaReceived October 23, 2002The kinetic and chemical mechanisms of AChE-catalyzed hydrolysis of short-chain thiocholine esters arerelatively well documented. Up to propanoylthiocholine (PrTCh) the chemical mechanism is general acid-basecatalysis by the active site catalytic triad. The chemical mechanism for the enzyme-catalyzed butyrylthio-choline(BuTCh) hydrolysis shifts to a parallel mechanism in which general base catalysis by E199 of directwater attack to the carbonyl carbon of the substrate. [Selwood, T., et al. J. Am. Chem. Soc. 1993, 115, 10477-10482] The long chain thiocholine esters such as hexanoylthiocholine (HexTCh), heptanoylthiocholine(HepTCh), and octanoylthiocholine (OcTCh) are hydrolyzed by electric eel acetylcholinesterase (AChE). Thekinetic parameters are determined to show that these compounds have a lower Michaelis constant than BuTChand the pH-rate profile showed that the mechanism is similar to that of BuTCh hydrolysis. The solvent isotopeeffect and proton inventory of AChE-catalyzed hydrolysis of HexTCh showed that one proton transfer isinvolved in the transition state of the acylation stage. The relationship between the dipole moment and theMichaelis constant of the long chain thiocholine esters showed that the dipole moment is the most importantfactor for the binding of a substrate to the enzyme active site.Key Words : Acetylcholinesterase, Hydrolysis, Thiocholine esters, Kinetic studies, Chemical mechanismIntroductionAcetylcholinesterase (AChE, EC 3.1.1.7) is a serinehydrolase that catalyzes the hydrolysis of the neurotrans-mitter, acetylcholine (ACh) to acetate and choline atneuromuscular synapses.