Microcalorimetric study of the inhibition of butyrylcholinesterase by paraoxon. - Université de Limoges Accéder directement au contenu
Article Dans Une Revue Analytical Biochemistry Année : 2009

Microcalorimetric study of the inhibition of butyrylcholinesterase by paraoxon.


The inhibition of horse serum butyrylcholinesterase (EC by the organophosphorus compound paraoxon (diethyl 4-nitrophenyl phosphate) was studied by flow microcalorimetry at 37 degrees C in Tris buffer (pH 7.5) using a modification of the kinetic model described by Stojan and coworkers [J. Stojan, V. Marcel, S. Estrada-Mondaca, A. Klaebe, P. Masson, D. Fournier, A putative kinetic model for substrate metabolisation by Drosophila acetylcholinesterase, FEBS Lett. 440 (1998) 85-88]. The reversible steps of the inhibition were studied in the mixing cell of the calorimeter, whereas the irreversible step was studied in the flow-through cell. A new pseudo-first-order approximation was developed to allow the kinetic analysis of inhibition progress curves in the presence of substrate when a significant amount of substrate is transformed. This approximation also allowed one to compute an analytical expression of the calorimetric curves using a gamma distribution to describe the impulse response of the calorimeter. Fitting models to data by nonlinear regression, with simulated annealing as a stochastic optimization method, allowed the determination of all kinetic parameters. It was found that paraoxon binds to both the enzyme and acyl-enzyme, but with weak affinities (K(i) = 0.123 mM and K'(i) = 5.5 mM). A slight activation was observed at the lowest paraoxon concentrations and was attributed to the binding of the substrate to the enzyme-inhibitor complex. The bimolecular inhibition rate constant k(i) = 2.8 x 10(4) M(-1) s(-1) was in agreement with previous studies. It is hoped that the methods developed in this work will contribute to extending the application range of microcalorimetry in the field of irreversible inhibitors.

Dates et versions

hal-00626897 , version 1 (27-09-2011)



Jean Debord, Michel Harel, Bernard Verneuil, Jean-Claude Bollinger, Thierry Dantoine. Microcalorimetric study of the inhibition of butyrylcholinesterase by paraoxon.. Analytical Biochemistry, 2009, 389 (2), pp.97-101. ⟨10.1016/j.ab.2009.03.041⟩. ⟨hal-00626897⟩
86 Consultations
0 Téléchargements



Gmail Facebook Twitter LinkedIn More