Article ID | Journal | Published Year | Pages | File Type |
---|---|---|---|---|
1982765 | Insect Biochemistry and Molecular Biology | 2009 | 8 Pages |
Abstract
Acetylcholinesterases (AChEs) and their genes from susceptible and resistant insects have been extensively studied to understand the molecular basis of target site insensitivity. Due to the existence of other resistance mechanisms, however, it can be problematic to correlate directly a mutation with the resistant phenotype. An alternative approach involves recombinant expression and characterization of highly purified wild-type and mutant AChEs, which serves as a reliable platform for studying structure-function relationships. We expressed the catalytic domain of Anopheles gambiae AChE1 (r-AgAChE1) using the baculovirus system and purified it 2,500-fold from the conditioned medium to near homogeneity. While KM's of r-AgAChE1 were comparable for ATC, AβMTC, PTC, and BTC, Vmax's were substantially different. The IC50's for eserine, carbaryl, paraoxon, BW284C51, malaoxon, and ethopropazine were 8.3, 72.5, 83.6, 199, 328, and 6.59 Ã 104 nM, respectively. We determined kinetic constants for inhibition of r-AgAChE1 by four of these compounds. The enzyme bound eserine or paraoxon stronger than carbaryl or malaoxon. Because the covalent modification of r-AgAChE1 by eserine occurred faster than that by the other compounds, eserine is more potent than paraoxon, carbaryl, and malaoxon. Furthermore, we found that choline inhibited r-AgAChE1, a phenomenon related to the enzyme activity decrease at high concentrations of acetylcholine.
Keywords
5,5-dithio-bis(2-nitrobenzoic acid)propionylthiocholine iodideBW284C51butyrylthiocholine iodideNi-NTABTCATCDTNBPTCAChEAcetylthiocholine iodideOrganophosphateAcetylcholinesterasesodium dodecyl sulfate-polyacrylamide gel electrophoresisSDS-PAGEOrganophosphorous compoundsCholinergic synapseMalariaInsecticide resistancenickel-nitrilotriacetic acidMosquitoCarbamatehigh performance liquid chromatographyHPLC
Related Topics
Life Sciences
Agricultural and Biological Sciences
Insect Science
Authors
Haobo Jiang, Siwei Liu, Picheng Zhao, Carey Pope,