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Figure 5.6 Stochastic properties of
chemoattractant receptor. (A) Schematic
drawing of single-molecule imaging of
Cy3
by measuring the time that elapsed between the
appearance and the disappearance of the
fluorescent spots. The line represents the fit of
the data to the sum of two exponential functions
with dissociation rates of 1.0 and 0.13 s 1 .
(D) Time course of receptor occupancy in
chemotaxing cells. The number of Cy3
cAMP bound to cAMP receptors. The basal
membrane of Dictyosteliumcells was observed by
total internal reflection fluorescence microscopy.
Cy3
-
cAMP molecules are visible as fluorescent
spots when they bind to the receptors. (B) Single-
molecule imaging of Cy3
-
cAMP
spots bound to the basal surface of the cells was
counted. The numbers of Cy3
-
cAMP bound to
Dictyostelium cells. Scale bar, 1
-
cAMP bound to
the anterior (black line) and the posterior (gray
line) halves of the cells exhibited fluctuations in
the receptor occupancy, as shown by the noisy
input signals.
-
m.
(C) Cumulative frequency histogram of the
lifetime of Cy3
m
-
cAMP spots. The lifetimes of
individual Cy3
-
cAMP molecules were obtained
membrane of living Dictyostelium cells at the single molecule level (Figure 5.6A
and B) [22]. The stochastic properties of the receptors were examined by determining
the lifetime of the cAMP - receptor complex and receptor occupancy in the living cells.
The lifetimes of the cAMP - receptor complexes exhibit an exponential distribution
(Figure 5.6C), meaning that ligand dissociation from the receptor is a random
process. The time series of receptor occupancy exhibits fluctuations with exponential
time correlations, revealing that ligand binding follows a Poisson distribution. Thus,
ligand binding takes place in a random manner and the input signals are therefore
noisy.
Figure 5.6D shows receptor occupancy in the cells moving toward the higher
concentration of Cy3 - cAMP. Receptor occupancy was sometimes inversely propor-
tional to the gradient of the chemoattractant. In our experimental system, only the
basal surface of the cells could be visualized, and hence the receptor occupancy shown
here does not represent the total input of chemotactic signals to the entire surface of
 
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