Acetylcholinesterases from the Disease Vectors Aedes aegypti and Anopheles gambiae: Functional Characterization and Comparisons with Vertebrate Orthologues

Abstract

Mosquitoes of the Anopheles (An.) and Aedes (Ae.) genus are principal vectors of human diseases including malaria, dengue and yellow fever. Insecticide-based vector control is an established and important way of preventing transmission of such infections. Currently used insecticides can efficiently control mosquito populations, but there are growing concerns about emerging resistance, off-target toxicity and their ability to alter ecosystems. A potential target for the development of insecticides with reduced off-target toxicity is the cholinergic enzyme acetylcholinesterase (AChE). Herein, we report cloning, baculoviral expression and functional characterization of the wild-type AChE genes (ace-1) from An. gambiae and Ae. aegypti, including a naturally occurring insecticide-resistant (G119S) mutant of An. gambiae. Using enzymatic digestion and liquid chromatography-tandem mass spectrometry we found that the secreted proteins were post-translationally modified. The Michaelis-Menten constants and turnover numbers of the mosquito enzymes were lower than those of the orthologous AChEs from Mus musculus and Homo sapiens. We also found that the G119S substitution reduced the turnover rate of substrates and the potency of selected covalent inhibitors. Furthermore, non-covalent inhibitors were less sensitive to the G119S substitution and differentiate the mosquito enzymes from corresponding vertebrate enzymes. Our findings indicate that it may be possible to develop selective non-covalent inhibitors that effectively target both the wild-type and insecticide resistant mutants of mosquito AChE.

Discovery of Species-selective and Resistance-breaking Anticholinesterase Insecticides for the Malaria Mosquito

“…Thus, it is to be expected that some substrate analog inhibitors of AChE (organophosphates, carbamates) may also show reduced sensitivity to G119S Ag AChE. Recently, Ag AChE (WT and G119S) and Aedes aegypti AChE (WT, Ae AChE) were also expressed in Sf9 cells [23]. Similar kinetic parameters ( K m , k cat ) were found for Ag AChE and Ae AChE.…”

Identification of Potential Inhibitors from Pyriproxyfen with Insecticidal Activity by Virtual Screening

“…aegypti , respectively; notably, the mosquito ( Ae. aegypti) and human acetylcholinesterase enzymes exhibit slightly increased sequence identity 48–49% [9,10,11]. This justifies several in vitro studies of acetylcholinesterase activity in order to confirm such enzymatic inhibition; as an example, Botas et al (2017) [10] carried out a study on the chemical composition, anticholinesterase activity, and nanoemulsions of limonene as a larvicidal agent for the control of Ae.…”

“…The insecticide pyriproxyfen (2-[1-methyl-2-(4-phenoxyphenoxy) ethoxy] pyridine) [9], as shown in Figure 1, is an aromatic compound, non-terpenoidal hormone, a potent suppressor of embryogenesis of insects at the adult stage, and belongs to the chemical group pyridyloxypropyl ether, synthesized and developed by Sumitomo Chemical Co., Ltd. in the 1990s [18,19,20].…”

Crystal structure of acetylcholinesterase catalytic subunits of the malaria vector Anopheles gambiae

“…After diligently considering the preferred glycans of yeast glycosylation system and observed Fo‐Fc and 2Fo‐Fc electro density maps, we modeled 2 N‐acetyl glucosamine molecules that link residue N220 of the structure. The glygosylation site is different from that reported previously, but the linkage is similar (Engdahl et al ., ).…”