Olfactory Receptor Proteins - Bioelectronic Nose: Integration of Biotechnology and Nanotechnology - page 62

reduced affinity for the odorant. At high ligand doses, and in absence of OBPs, the
OR dimer could then bind two odorant molecules, one on each protomer, and this
would inactivate the receptors. Yet, in presence of OBPs and at high odorant doses,
OBPs binding to the OR dimer at an allosteric site would prevent the binding of a
second odorant molecule and would thus preserve OR activity. Such “multi-state”
models have already been reported for other GPCRs [ 63 ], in which the activity de-
pends on the occupation rate of the various sites on the dimers.
This negative modulation of OR activity by odorants themselves is essential
to take into account when searching to identify OR odorant ligands or to detect
odorants using biosensors carrying ORs.

3.6 Use of Olfactory Receptor Protein in

Bioelectronic Noses

The animal olfactory system, with its ability to identify and discriminate thousands
of odorant compounds with very low thresholds (10 −11 -10 −17 M for some odorants
in humans or in dogs, [ 64 , 65 ]), is worth mimicking to engineer bioelectronic noses.
Using ORs as basic sensing elements instead of chemical sensors in electronic nos-
es allows researchers to benefit from the naturally optimized molecular recognition
of odorants to develop these new devices. ORs first have to be expressed in heter-
ologous systems, which is not yet an easy task (see the next chapters for the various
expression systems considered). Then, several procedures can be used depending
on the method considered for measuring their functional response, in order to im-
prove the technical performances of the devices.
ORs can be purified in the presence of surfactants, or prepared as membrane
fractions, micelles, nanodisks, nanoliposomes or nanovesicles, then specifically
grafted on functionalized surfaces, including gold electrodes or carbon or poly-
mer nanotubes. Their functional response can then be monitored by physical or
biophysical measurements, at submillimetric to micrometric scales. Detection lim-
its and odorants discrimination can in some cases reach the femtomolar (fM), an
intrinsic property of the ORs themselves.
Olfactory receptors carrying a His tag can be produced in a baculovirus/insect
cells system [ 66 ] or in Escherichia coli [ 67 ], and affinity-purified using nickel-
magnetic beads. The response of these ORs prepared either as digitonin micelles, or
as soluble nanodiscs, and deposited on nickel-nitrilotriacetic acid (Ni-NTA) carbon
nanotubes (CNTs) can be followed by a change of conductivity measured by the
CNT transistor [ 66 ]. Alternately, purified ORs are deposited on conducting poly-
mers nanotubes (CPNTs) [ 68 ], or on single-wall functionalized carbon nanotubes
[ 67 ]. With CPNTs deposited on interdigitated microelectrodes, the resulting resis-
tance varies linearly between the detection limit of 0,02 ppt and 2 ppm.
Starting from HEK293 cells, nanovesicles with a diameter of 100-200 nm car-
rying ORs can be prepared using cytochalasin [ 69 , 70 ], and directly attached to
carbon nanotubes (swCNT). In this case, exposition of the nanovesicles to the