Title: The influence of peripheral site ligands on the reaction of symmetric and chiral organophosphates with wildtype and mutant acetylcholinesterases Radic Z, Taylor P Ref: Chemico-Biological Interactions, 119-120:111, 1999 : PubMed
The rates of inhibition of mouse acetylcholinesterase (AChE) (EC 3.1.1.7) by paraoxon, haloxon, DDVP, and enantiomers of neutral alkyl methylphosphonyl thioates and cationic alkyl methylphosphonyl thiocholines were measured in the presence and absence of AChE peripheral site inhibitors: gallamine, D-tubocurarine, propidium, atropine and derivatives of coumarin. All ligands, except the coumarins, at submillimolar concentrations enhanced the rates of inhibition by neutral organophosphorus compounds (OPs) while inhibition rates by cationic OPs were slowed down. When peripheral site ligand concentrations extended to millimolar, the extent of the enhancement decreased creating a bell shaped activation profile. Analysis of inhibition by DDVP and haloxon revealed that peripheral site inhibitors increased the second order reaction rates by increasing maximal rates of phosphylation.
Title: Role of the peripheral anionic site on acetylcholinesterase: inhibition by substrates and coumarin derivatives Radic Z, Reiner E, Taylor P Ref: Molecular Pharmacology, 39:98, 1991 : PubMed
Propidium has been demonstrated in previous studies to be a selective ligand for the peripheral anionic site on acetylcholinesterase (EC 3.1.1.7). Its association with this site can be advantageously monitored by direct fluorescent titration. We have measured the ability of acetylcholine, acetylthiocholine, haloxon [di-(2-chloroethyl)3-chloro-4-methylcoumarin-7-ylphosphate] , and a coumarin derivative (3-chloro-7-hydroxy-4-methylcoumarin) to dissociate propidium from the peripheral anionic site of Torpedo californica acetylcholinesterase. Measurements were made by back-titration of propidium after complete inhibition of the active center with diisopropylfluorophosphate. Both acetylcholine and acetylthiocholine show substrate inhibition at high substrate concentrations. The concentrations required for occupation of the peripheral site, as ascertained by competition with propidium, correlated well with the concentration dependence for the kinetics of substrate inhibition. These observations are consistent with substrate inhibition being due to binding of acetylcholine or acetylthiocholine at a peripheral anionic site. Displacement of propidium by haloxon and coumarin indicated that these inhibitors also bind to the peripheral anionic site. The dissociation constants ascertained from peripheral site occupation are in agreement with the constants obtained from inhibition kinetics. Evidence is presented that competition with propidium obtained by direct fluorescence titrations, when combined with inhibition kinetics, provides a more reliable means for ascertaining site selectivity of various inhibitors than does a kinetic analysis alone.
