Characterization of a Pheromone-Degrading Enzyme from De Pest Moth

Characterization of a pheromone-degrading enzyme from de pest moth

Lobesia botrana

Our group deals with different aspects related with new insect pheromones, from structural characterization and synthesis to the establishment of the attractant activity in the laboratory and in the field. We are also engaged in the development of new alternative and non-contaminant methods of pest control, based on inhibition of the enzymes involved in the degradation of pheromone molecules in the insect’s antennae.

To study the interaction inhibitor-active center of the enzyme from a molecular point of view, we have started a collaboration with Drs. Emmanuelle Jacquin-Joly and Martine Maibeche (Station de Phytopharmacie, INRA, Versailles and University Pierre et Marie Curie, Paris) to study cloning, expression and molecular characterization of the antennal esterases. Up to now only few esterases of insects responsible for the degradation of the pheromone has been isolated, cloned and expressed. In a first stage, we will focus on the localization of the specific esterase(s) involved in the pheromone metabolism by electrophoretic techniques under non-denaturing conditions (PAGE) through comparison of the esterases present in different tissues of males and females. Later on, after extracting the RNA total through homogenization of the antennae we will proceed to obtain the cDNAs encoding the specific esterases of the catabolism through a PCR (Polymerase Chain Reaction) strategy. To do this, degenerate primers will be designed in base to sequences observed in esterases of other insects already cloned. From the complete sequence of the gene of interest, we will try to obtain the recombinant protein through expression in Escherichia coli, or better through expression in insect cell cultures using a baculovirus as expression vector. The activity of the recombinant and native enzyme will be compared through kinetic studies (Km, Vmax) of the pheromone degradation, including substrate specificity (Kcat/Km).

In conclusion, the object of this work is the molecular characterization, cloning and expression of the antennal esterases of L. botrana in order to get acceptable amounts of pure enzyme for future research.

To develop this project, it is necessary that the candidate should have experience in

A)Protein electrophoresis

B)RNA extraction and manipulation

C)RT-PCR (reverse Transcription-Polymerase Chain Reaction

D)cDNA cloning

Correspondence: Dra. Gloria Rosell.Unit of Medicinal Chemistry (Associated to CSIC).

Faculty of Pharmacy, University of Barcelona. Av. Diagonal, s/n.08028 Barcelona; e-mail:;