Acetylcholinesterase from Atta Sexdens (leaf-cutter ants): development of a system to expression, purification and functional characterization

Abstract

Leaf-cutting ants, especially species of the Atta and Acromyrmex genus, are known for their destructive power of a large number of plant species and the economic damage caused to agriculture. The most widely used insecticides target the inhibition of the enzyme acetylcholinesterase - AChE - (EC 3.1.1.7 ). Although widely used, these insecticides also inhibit AChE animals (including human) which causes serious environmental and health problems. This is because the catalytic site of insect and human AChE has a high similarity and insecticides that are inhibitors bind in this region of the molecule. Recent studies show the presence of a cysteine near catalytic site that is conserved in all insects but absent in mammals, which can be an interesting target for more specific and environmentally safe agricultural pesticides and herbicides. The relevance of this analysis is very important since there are possibilities, by means of techniques of rational design of molecules of the same or search on natural products, and specific create a fully effective compound, and , furthermore, environmentally safe .Thus, the aim of this project is to express AchE from A. sexdens in soluble and active form for use in assays in search inhibitors and in structural study. To achieve this goal, the gene encoding AchE will be cloned into vector suitable for expression in P. pastoris and the expressed enzyme will be purified and characterized enzymatically. The pure enzyme will be used in crystallization trials for future structural studies and will also be immobilized to optimize the search for inhibitors.