Evolution and properties of acetylcholinesterase during developmental stages of dengue vector (Aedes aegypti)

The screening of acetylcholinesterase (AChE) (E.C.3.1.1.7.) during developmental stages, eggs, larvae, pupa and adults of Aedes aegypti was detected. Chromatography on a Sephacryl S-200 column revealed the existence of two forms designated as AChE1 and AChE2. Different expression patterns of AChE1 and AChE2 activities were detected. The highest expression of AChE1 and AChE2 were detected at the egg and adult, respectively. The native molecular weights of AChE1 and AChE2 from Ae. aegypti were estimated to be > 2,000 and 100 kDa, respectively. The properties of AChE1 and AChE2 from Ae. aegypti 36h-old eggs were studied. The substrate specificity of enzymes showed that the relative activity % for butyrylthiocholine iodide was 9.4 and 11.6 for AChE1 and AChE2 as compared to acetylthiocholine iodide (AcSChI), respectively. The Km values for Ae. aegypti AChE1 and AChE2 were 0.256 and 0.288 mM AcSChI, respectively. Both enzymes had the same temperature and pH optima at 40˚C and 7.5, respectively. AChE1 was thermal stable up to 40˚C, while AChE2 lost 20% of its activity. All the metal examined caused partial inhibitory effect on the two enzymes, except Co+2 caused complete inhibition. While PMSF, 1,10-phenanthroline, trypsin inhibitor and EDTA caused partial inhibitory effect on both enzymes, the activities were completely lost by p-HMB. The type of inhibition, bimolecular rate constant (Ki) and Ki ratio AChE1/AChE2 for organophosphate or carbamate as inhibitors were detected. AChE1 was 0.9-, 0.69- and 0.48-fold more sensitive to inhibition by methomyl, chlorpyrifos-methyl and pirimiphos-methyl, and 4.3-, 1.9- and 1.36-fold less sensitive to inhibition by eserine, malathion and fenitrothion than that of AChE2, respectively. These results could support vector control program by choosing the right pesticide which specifically inhibits any of the molecular forms of AChE’s at the right developmental stage. In addition, the obtained results provides important biochemical characteristics of the enzyme AChE from the dengue vector Ae. aegypti, the target of neuropesticides such as organophosphorus, carbamates and pyrothroid compounds.