Approaches towards understanding insect odorant receptor structure and function

The ability to smell volatile compounds is essential for all insects, being involved in locating food, hosts, enemies and potential mates. In insects olfaction is largely mediated by members of a novel family of ligand-gated cation channels. While these receptors have seven transmembrane regions, their N-terminus is cytoplasmic, unlike G protein-coupled receptors that have an extracellular N-terminus. The active complex contains a specific ligand binding subunit and the ubiquitous non-canonical subunit Orco (previously Or83b), which is absolutely required for function. We are using are range of approaches to understand the structure and function of odorant receptors from the model insect, Drosophila melanogaster, with the aims of developing novel insect control strategies and olfactory biosensors. Intermolecular Förster Resonance Energy Transfer (FRET) using recombinant fluorescent protein fusions is being used to investigate homo- or hetero-dimerisation or oligomerisation of receptor subunits, while changes in both inter- and intra-molecular FRET upon the addition of ligand is being used to infer activation mechanisms. We are also attempting Surface Plasmon Resonance to detect receptor subunit interaction and ligand binding onto immobilised recombinant receptor. Finally, comparison of chimeras of orthologous versions of ligand binding receptors is being used to identify regions within receptors that are involved in specificity and therefore ligand binding. Using this approach a region within the sixth transmembrane region of the receptor Or22a has been identified that is involved in the ability to detect 2-heptanone.