[Medical diagnosis by breath analysis: odor sensors].

An odor sensor is a device for detecting target odors within a mixture, used in many fields including medical diagnosis. Electronic noses are networks of olfactory sensors, consisting of a surface whose properties are modified in the presence of odors, coupled with a measurement system. Their olfactory signature is analyzed in comparison with databases.

Immunochemical strategy for quantification of G-coupled olfactory receptor proteins on natural nanovesicles.

Cell membrane proteins are involved in a variety of biochemical pathways and therefore constitute important targets for therapy and development of new drugs. Bioanalytical platforms and binding assays using these membrane protein receptors for drug screening or diagnostic require the construction of well-characterized liposome and lipid bilayer arrays that act as support to prevent protein denaturation during biochip processing. Quantification of the protein receptors in the lipid membrane arrays is a key issue in order to produce reproducible and well-characterized chips.

A novel platform based on immobilized histidine tagged olfactory receptors, for the amperometric detection of an odorant molecule characteristic of boar taint.

We report a dose-dependent detection of androstenone in solution, as one of the boar taint compounds, based on related OR7D4 olfactory receptors immobilized on a gold electrode through their 6-His tag and NTA-copper complex, as visualized through fluorescence microscopy. Square wave voltammetry (SWV) is for the first time, the method used to monitor the olfactory receptor/odorant recognition process. The relative variation of the Cu(I)-Cu(II) current peak increases linearly versus log (concentration of androstenone) from 10(-14)M to 10(-4)M, in buffer solution.

Mammalian olfactory receptors: molecular mechanisms of odorant detection, 3D-modeling, and structure-activity relationships.

This chapter describes the main characteristics of olfactory receptor (OR) genes of vertebrates, including generation of this large multigenic family and pseudogenization. OR genes are compared in relation to evolution and among species. OR gene structure and selection of a given gene for expression in an olfactory sensory neuron (OSN) are tackled.

Promotion of cancer cell invasiveness and metastasis emergence caused by olfactory receptor stimulation.

Olfactory receptors (ORs) are expressed in the olfactory epithelium, where they detect odorants, but also in other tissues with additional functions. Some ORs are even overexpressed in tumor cells. In this study, we identified ORs expressed in enterochromaffin tumor cells by RT-PCR, showing that single cells can co-express several ORs.

Force measurements on natural membrane nanovesicles reveal a composition-independent, high Young's modulus.

Mechanical properties of nano-sized vesicles made up of natural membranes are crucial to the development of stable, biocompatible nanocontainers with enhanced functional, recognition and sensing capabilities. Here we measure and compare the mechanical properties of plasma and inner membrane nanovesicles ∼80 nm in diameter obtained from disrupted yeast Saccharomyces cerevisiae cells. We provide evidence of a highly deformable behaviour for these vesicles, able to support repeated wall-to-wall compressions without irreversible deformations, accompanied by a noticeably high Young's modulus (∼300 MPa) compared to that obtained for reconstituted artificial liposomes of similar size and approaching that of some virus particles.

Early survival factor deprivation in the olfactory epithelium enhances activity-driven survival.

The neuronal olfactory epithelium undergoes permanent renewal because of environmental aggression. This renewal is partly regulated by factors modulating the level of neuronal apoptosis. Among them, we had previously characterized endothelin as neuroprotective.

Deciphering activation of olfactory receptors using heterologous expression in Saccharomyces cerevisiae and bioluminescence resonance energy transfer.

Hetero- and homo-oligomerization of G protein-coupled receptors (GPCRs) has been addressed in the past years using various approaches such as co-immunoprecipitation, fluorescence resonance energy transfer and bioluminescence resonance energy transfer (BRET). Here, we report the methodological details from a previously published study to investigate the relationships between oligomerization and activation states of olfactory receptors (ORs). This methodology combines heterologous expression of ORs in Saccharomyces cerevisiae and BRET assays on membrane fractions, in particular, upon odorant stimulation.

Modeling of mammalian olfactory receptors and docking of odorants.

Olfactory receptors (ORs) belong to the superfamily of G protein-coupled receptors (GPCRs), the second largest class of genes after those related to immunity, and account for about 3 % of mammalian genomes. ORs are present in all multicellular organisms and represent more than half the GPCRs in mammalian species (e.g.

Automatic modeling of mammalian olfactory receptors and docking of odorants.

We present a procedure that (i) automates the homology modeling of mammalian olfactory receptors (ORs) based on the six three-dimensional (3D) structures of G protein-coupled receptors (GPCRs) available so far and (ii) performs the docking of odorants on these models, using the concept of colony energy to score the complexes. ORs exhibit low-sequence similarities with other GPCR and current alignment methods often fail to provide a reliable alignment. Here, we use a fold recognition technique to obtain a robust initial alignment.

Relationship between homo-oligomerization of a mammalian olfactory receptor and its activation state demonstrated by bioluminescence resonance energy transfer.

G-protein-coupled receptor homo-oligomerization has been increasingly reported. However, little is known regarding the relationship between activation of the receptor and its association/conformational states. The mammalian olfactory receptors (ORs) belong to the G protein-coupled receptor superfamily.

On a chip demonstration of a functional role for Odorant Binding Protein in the preservation of olfactory receptor activity at high odorant concentration.

The molecular mechanisms underlying odorant detection have been investigated using the chip based SPR technique by focusing on the dynamic interactions between transmembrane Olfactory Receptor OR1740, odorant ligands and soluble Odorant-Binding Protein (OBP-1F). The OR1740 present in the lipid bilayer of nanosomes derived from transformed yeasts specifically bound OBP-1F. The receptor preferential odorant ligand helional released bound OBP-1F from the OR-OBP complex, while unrelated odorants failed to do so.

A new concept of olfactory biosensor based on interdigitated microelectrodes and immobilized yeasts expressing the human receptor OR17-40.

This work shows the feasibility of an olfactory biosensor based on the immobilization of Saccharomyces cerevisiae yeast cells genetically modified to express the human olfactory receptor OR17-40 onto interdigitated microconductometric electrodes. This olfactory biosensor has been applied to the detection of its specific odorant (helional) with a high sensitivity (threshold 10(-14) M). In contrast, no significant response was observed using a non-specific odorant (heptanal), which suggests a good selectivity.

Gold surface functionalization and patterning for specific immobilization of olfactory receptors carried by nanosomes.

There is substantial interest in engineering solid supports to achieve functional immobilization of membrane receptors both for investigation of their biological function and for the development of novel biosensors. Three simple and practical strategies for immobilization of a human olfactory receptor carried by nanosomes are presented. The basis of the functionalization of solid gold surfaces is a self-assembled monolayer (SAM) containing biotinyl groups.

A novel detection strategy for odorant molecules based on controlled bioengineering of rat olfactory receptor I7.

In this study, we report a dose-dependent detection of odorant molecules in solution by rat olfactory receptor I7 (OR I7) in its membrane fraction. The OR I7 is immobilized on a gold electrode by multilayer bioengineering based on a mixed self-assembled monolayer and biotin/avidin system, which allows for a well-controlled immobilization of the bioreceptor within its lipid environment. The odorant detection is electronically performed in a quantitative manner by electrochemical impedance spectroscopy (EIS) measurements on samples and controls.

Immobilization of rhodopsin on a self-assembled multilayer and its specific detection by electrochemical impedance spectroscopy.

Rhodopsin, the G protein-coupled receptor (GPCR) which mediates the sense of vision, was prepared from calf eyes and used as receptor enriched membrane fraction. In this study it was immobilized onto gold electrode by two different techniques: Langmuir-Blodgett (LB) and a strategy based on a self-assembled multilayer. We demonstrated that Langmuir and LB films of rhodopsin are not stable.

Immobilization of native membrane-bound rhodopsin on biosensor surfaces.

In this paper, we evaluated the grafting of G-protein-coupled receptors (GPCRs) onto functionalized surfaces, which is a primary requirement to elaborate receptor-based biosensors, or to develop novel GPCR assays. Bovine rhodopsin, a prototypical GPCR, was used in the form of receptor-enriched membrane fraction. Quantitative immobilization of the membrane-bound rhodopsin either non-specifically on a carboxylated dextran surface grafted with long alkyl groups, or specifically on a surface coated with anti-rhodopsin antibody was demonstrated by surface plasmon resonance.

Functional expression of olfactory receptors in yeast and development of a bioassay for odorant screening.

The functional expression of olfactory receptors (ORs) is a primary requirement to examine the molecular mechanisms of odorant perception and coding. Functional expression of the rat I7 OR and its trafficking to the plasma membrane was achieved under optimized experimental conditions in the budding yeast Saccharomyces cerevisiae. The membrane expression of the receptor was shown by Western blotting and immunolocalization methods.

Engineered yeasts as reporter systems for odorant detection.

The functional expression of olfactory receptors (ORs) is a primary requirement to utilize olfactory detection systems. We have taken advantage of the functional similarities between signal transduction cascades in the budding yeast Saccharomyces cerevisiae and mammalian cells. The yeast pheromone response pathway has been adapted to allow ligand-dependent signaling of heterologous expressed G-protein coupled receptors (GPCRs) via mammalian or chimeric yeast/mammalian Galpha proteins.

Ligand-specific dose-response of heterologously expressed olfactory receptors.

Primary olfactory neuronal cultures exposed to odorant stimulation have previously exhibited concentration-related effects in terms of intracellular cAMP levels and adenylate cyclase activity [Ronnett, G.V., Parfitt, D.

Rescue of intracellularly trapped lutropin receptor exodomain by endodomain and reconstitution of a functional membrane receptor: interaction between exo- and endodomains.

The lutropin receptor consists of an extracellular N-terminal half and a membrane-associated C-terminal half. hCG initially binds the exodomain with a high affinity and the resulting complex is thought to interact with the endodomain through a secondary contact generating a hormonal signal. Therefore, the exodomain and endodomain are likely to associate directly or indirectly with each other, but lack of fruitful materials and technology has hampered knowledge about their physical relationship and contact sites.