The Molecular Basis for Ligand Specificity in a Mouse Olfactory Receptor

Abstract

Sequence differences between members of the mouse olfactory receptor MOR42 subfamily (MOR42-3 and MOR42-1) are likely to be the basis for variation in ligand binding preference among these receptors. We investigated the specificity of MOR42-3 for a variety of dicarboxylic acids. We used site-directed mutagenesis, guided by homology modeling and ligand docking studies, to locate functionally important residues. Receptors were expressed in Xenopus oocytes and assayed using high throughput electrophysiology. The importance of the Val-113 residue, located deep within the receptor, was analyzed in the context of interhelical interactions. We also screened additional residues predicted to be involved in ligand binding site, based on comparison of ortholog/paralog pairs from the mouse and human olfactory receptor genomes (Man, O., Gilad, Y., and Lancet, D. (2004) Protein Sci. 13, 240 -254). A network of 8 residues in transmembrane domains III, V, and VI was identified.These residues form part of the ligand binding pocket of MOR42-3. C12 dicarboxylic acid did not activate the receptor in our functional assay, yet our docking simulations predicted its binding site in MOR42-3. Binding without activation implied that C12 dicarboxylic acid might act as an antagonist. In our functional assay, C12 dicarboxylic acid did indeed act as an antagonist of MOR42-3, in agreement with molecular docking studies. Our results demonstrate a powerful approach based on the synergy between computational predictions and physiological assays.

Odorant Receptor Polymorphisms and Natural Variation in Olfactory Behavior in Drosophila melanogaster

“…Amino acid substitutions in vertebrate chemosensory receptors can result in changes in ligandbinding affinity and confer differences in odorant sensitivity (Krautwurst et al 1998;Feinstein and Mombaerts 2004;Reed et al 2004;Nie et al 2005;Abaffy et al 2007;Keller et al 2007). Thus, it is of particular interest that Or43a harbors an isoleucine to methionine substitution that contributes to variation in responsiveness to benzaldehyde.…”

Olfactory receptor 544 reduces adiposity by steering fuel preference toward fats

“…Azelaic acid (AzA) and suberic acid are known Olfr544 ligands (10,11). We examined the effect of Olfr544 in Hana3A cells transfected with N-terminal Lucy-FLAG-tagged Olfr544.…”

Modeling of mammalian olfactory receptors and docking of odorants

“…For ORs, this point might not be too critical, since odorants are usually low-molecular-weight organic compounds and therefore relatively small compared to the ligands of other GPCRs. A number of OR models in which odorants were docked and which were tested by sitedirected mutations have revealed that odorants dock deep into the pocket (Abaffy et al 2007;Katada et al 2005;Launay et al 2012) and do not interact with ECL2.…”

“…Using site-directed mutagenesis and functional expression the authors show that the orientation of odorants within a homo logy modelingderived binding pocket of olfactory receptor orthologs is defined by evolutionary conserved amino acid positions. mOR42-3, mOR42-1 2.8 Å bovine rhodopsin 3D structure Abaffy et al 2007 Investigation of the specificity of mOR42-3 for dicarboxylic acids. Use of site-directed mutagenesis to locate functionally important residues.…”