title: Kinetic insight into the mechanism of cholinesterasterase inhibition by aflatoxin B1 to develop biosensors.
abstract: In this paper, the inhibition effect of aflatoxin B1 on different species of cholinesterases was investigated to unravel action mechanism. The inhibition curves of several cholinesterase mutants (obtained by spectrophotometric measurements of enzyme activity, pS curves) were analyzed. They showed that this toxin reversibly inhibits cholinesterases by binding to a peripheral site located at the entrance of the active site gorge without entering inside the site. Electric eel enzyme revealed the highest inhibition extent with a binding constant estimated to 0.35 microM. This binding prevents the entrance of substrate en route to the catalytic site and also decreases chemical steps of the reaction at the catalytic site: acetylation is reduced to the half and deacetylation is reduced to the third. Electric eel acetylcholinesterase was used to settle an amperometric biosensor. The best detection was obtained by using 0.3 mU enzyme on the electrode and 0.5mM ATCh in the solution. The limit of detection was 3 microM corresponding to 20% inhibition.
authors.affiliation: Université de Toulouse, IPBS-CNRS 205 route de Narbonne, Toulouse, France.
authors.forename: Tamara
authors.initials: T
authors.lastname: Hansmann
authors.forename: Benoît
authors.initials: B
authors.lastname: Sanson
authors.forename: Jure
authors.initials: J
authors.lastname: Stojan
authors.forename: Martin
authors.initials: M
authors.lastname: Weik
authors.forename: Jean-Louis
authors.initials: JL
authors.lastname: Marty
authors.forename: Didier
authors.initials: D
authors.lastname: Fournier
mesh_terms: D000110:Acetylcholinesterase / Q000737:chemistry*; D016604:Aflatoxin B1 / Q000032:analysis* / Q000037:antagonists & inhibitors*; D015374:Biosensing Techniques / Q000295:instrumentation* / Q000379:methods; D002800:Cholinesterase Inhibitors / Q000032:analysis / Q000737:chemistry*; D003198:Computer Simulation; D017076:Computer-Aided Design; D004563:Electrochemistry / Q000295:instrumentation*; D004566:Electrodes; D004789:Enzyme Activation; D004867:Equipment Design; D019544:Equipment Failure Analysis; D007700:Kinetics; D008956:Models, Chemical; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity
chemical_list: D002800:Cholinesterase Inhibitors; D016604:Aflatoxin B1; D000110:Acetylcholinesterase