Biomedical Engineering Reference
In-Depth Information
OR-odorant interactions or indirect detection via signal transduction by the binding
OR-odorant in living cell. SPR techniques are implemented for real-time monitor-
ing of step-by-step surface immobilization to test the functional response of an im-
mobilized OR [
103
]. ORs were immobilized onto a gold surface functionalized by
mixed self-assembled monolayers and biotin/neurtravidin. The study showed that
the specific immobilization of capturing the OR via an antibody yields a uniform
orientation of OR protein onto the surface while keeping the receptor activity pre-
served by a binding assay. Vidic et al. reported that the rORI7 and hOR17-40 could
be functionally expressed in
S. cerevisiae
[
15
]. They also developed yeast-derived
nanosomes with OR protein-grafted SPR technique to quantitatively evaluate OR
stimulation by an odorant (see Fig.
11.9
).
In another example, Benilova et al. described OR-carrying nanosomes based
olfactory biosensor using SPR [
104
]. The hOR17-40 was heterologously co-ex-
pressed with G
αolf
protein in yeast, and nanosomes were prepared and then specif-
ically immobilized. The result showed the bell-shaped response profile with two
maximum (1 nM and 1 μM) observed for the helional, which is a specific odor-
ant. Interestingly, the molecular mechanisms by focusing on dynamic interactions
between OR, odorant binding protein (OBP) and odorant was investigated using
the SPR technique [
14
]. OBPs can bind and solubilize volatile odorants, facili-
tating their diffusion through the mucus barrier towards ORs. The high-affinity
binding of a porcine OBP to a human OR has been demonstrated, suggesting that
a specific OR-OBP association may occur in the absence of any odorant [
105
].
Results show that SPR-based assay enables the direct demonstration of interac-
tions between the three components involved in the initial step of olfactory signal
transduction.
The SPR technique was used for the analysis of interaction between living
olfactory cells and odorants. This technique was applied to the cell-based mea-
surement of odorant molecules [
13
,
102
]. ODR-10, with the help of the rho-tag
import sequence, the OR protein of the nematode
C. elegans
, was expressed on
the surface of the HEK-293 cell and cultured in Au film. Exposure of the cells
to diacetyl, an odorant molecule specific to the ODR-10, induced a SPR signal.
The ODR-10 on the
C. elegans
neuron is coupled to an IP
3
pathway, which leads
to the release of Ca
2+
from the intracellular stores such as ER [
106
,
107
]. This
intracellular signaling through the binding of odorant molecules to OR can make
SPR signals. In other example, Real-time monitoring of odorant-induced cellu-
lar reaction via signal transduction was demonstrated using an SPR system [
12
].
The key features are the intracellular signal transduction triggered by the specific
binding of odorants to the ORs and the SPR response generated by the intracel-
lular change. The SPR response to the odorant molecules was quantitative and
selective. It was linearly dependent on the odorant concentration and the response
signal was much lower than other odorant molecules sharing similar chemical
structures and properties.
Search WWH ::
Custom Search