Investigating the molecular basis of ligand binding to and activation of olfactory receptors in living cells

The olfactory system is a highly specialized chemodetector capable of discriminating between thousands of volatile chemical substances. This enormous capacity of chemical recognition is based on a large family of olfactory receptors (ORs). Despite the fact that quite a number of putative ORs have been cloned, only limited progress has been made in functional expression of these receptors in heterologous cells. In order to functionally express odorant receptors we have established a suitable expression system in HEK 293 cells. The co-expression of the Gαq protein together with odorant receptors allowed the study of OR activation by calcium imaging. To analyze the structural properties of odorant molecules essential for OR5 and hOR17-4 binding and activation, we developed an efficient functional assay based on optical read-out of odorant responses by intracellular calcium imaging on living cells. Investigation of the ligand specificity of the two ORs derived from different species, but exhibiting significant homology on amino acid sequence level, showed a high degree of similarity in their responsive odorant molecule spectrum. A comparison of the molecular properties of odorant compounds crucial for OR5 and hOR17-4 binding and activation revealed a classification in two different structural groups. The first one comprises aromatic hydrocarbons with variable structures and sizes and the second group consists of aliphatic odorant molecules which are mainly aldehydes with variable carbon chain lengths. To elucidate further the function of ORs, the computational modeling to predict the binding site of OR5 and the docking of the small, hydrophobic odorant molecule, safrole were determined. Our findings show that the safrole binding site in OR5 involves the helical domains TM3, TM5 and TM6, which is consistent with earlier computational studies on rat I7 and mouse OR-EG receptors. The key amino acid residues appear to be hydrophobic and the van der Waals forces are implicated in safrole binding to the OR5 receptor. The mutations of the amino acid residues predicted by safrole docking analyses, Val202, Phe251 and Leu258 to Ala, indicate that these residues play an important role in OR5 activation.

Vogel, Horst
Lausanne, EPFL
Other identifiers:
urn: urn:nbn:ch:bel-epfl-thesis3668-1

Note: The status of this file is: EPFL only

 Record created 2006-09-20, last modified 2018-03-17

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