Non-productive binding of butyryl(thio)choline in the active site of vertebrate acetylcholinesterase.
The kinetic behavior of cholinesterases is unconventional. While their activities are higher than expected by classical Michaelis-Menten reaction mechanisms, at intermediate substrate concentrations they show strong inhibition by excess of substrate. To date, the main explanations used for all of their kinetic peculiarities include hindrance of product exit, entropically improved water orientation by a second substrate molecule, and complete blockade of the fully occupied active site. However, with the hydrolysis of butyryl(thio)choline by vertebrate acetylcholinesterase, there are time-dependent and substrate-concentration-dependent decreases in catalytic activity. As the substrate depletion results in the expected downwardly concave shape of the progress curves for product formation at low substrate concentrations, this cannot be the reason for the bending of the linear progress curves at higher substrate concentrations. A good theoretical and practical explanation was reached by including the time-dependent appearance of a non-productive enzyme-substrate complex in the reaction scheme. The slow establishment of this complex appears to be a rare occurrence of incorrect substrate orientation at the bottom of the active site, with this blocked by a second substrate molecule.
Stojan
Jure
J
Institute of Biochemistry, Faculty of Medicine, University of Ljubljana, Vrazov Trg 2, 1000 Ljubljana, Slovenia. stojan@mf.uni-lj.si
D000110:Acetylcholinesterase / Q000737:chemistry* / Q000378:metabolism*; D000122:Acetylthiocholine / Q000378:metabolism; D000818:Animals; D002092:Butyrylthiocholine / Q000378:metabolism* / Q000494:pharmacology; D020134:Catalytic Domain; D002800:Cholinesterase Inhibitors / Q000378:metabolism / Q000494:pharmacology; D006868:Hydrolysis; D007700:Kinetics; D011485:Protein Binding; D014101:Torpedo
D002800:Cholinesterase Inhibitors; D000122:Acetylthiocholine; D002092:Butyrylthiocholine; D000110:Acetylcholinesterase