Biomedical Engineering Reference
In-Depth Information
further aggravated by the use of the peak amplitude as an indicator of the intensity of the
underlying photoelectric process. This latter pitfall can be made apparent by referring to a
schematic diagram shown in Figure 15.7. Both
B
1
and
B
2
are shown to have a biexponential
decay curve, as predicted by the equivalent circuit. The composite signal (
B
1
B
2
), which
can be observed at neutral pH and room temperature, is the algebraic sum of
B
1
and
B
2
.
Because of the partial overlap of the decay of
B
1
and the rise of
B
2
, inhibition of
B
2
alone
diminishes the negative peak but increases the positive peak, giving rise to the apparent
effect of a concurrent enhancement of
B
1
. In data reported by Drachev et al. [56], inhibition
of
B
2
by low pH had been partially attributed to an apparent enhancement of
B
1
, which did
not actually happen. This misinterpretation further conceals the already inconspicuous pH
dependence of
B
2
under open-circuit conditions. A similar problem also appears in the ERP
elicited from a reconstituted rhodopsin membrane and can be similarly explained [54].
What then is the molecular mechanism for
B
1
and for
B
2
?
B
1
and
B
2
respond very dif-
ferently to the manipulations of the aqueous phases.
B
2
is highly sensitive to pH [59], to
electrolyte composition [60,65], to proton-deuterium exchange [59], to chemical modifi-
cation of the membrane surface [62], and to site-directed mutagenesis [63]. In contrast,
B
1
is insensitive to all these treatments. These differences suggest that
B
1
is generated by
molecular processes occurring deep inside the hydrophobic portion of bR, whereas
B
2
is
generated at the hydrophilic domain at the membrane-water interface. In other words,
B
1
is generated by the OD mechanism whereas
B
2
is generated by the ICT mechanism, most
likely interfacial proton transfer. This assignment is further supported by an experiment
to be described next. In view of the drastically different effects of temperature, pH, pro-
ton-deuterium exchange, chemical modification, and amino-acid substitution, on the two
components, the physical methods of decomposition must be a clean one. Ad hoc mod-
els found in the literature usually could not achieve such a sweeping consistency.
A lingering question remains: why is
B
2
absent in an ML film? The answer to this ques-
tion is provided by a “Q-tip” experiment carried out by Michaile and Hong [61]. In the
“Q-tip” experiment, an ML film can be converted into a TM film by stripping the multiple
layers with a cotton swab (Q-tips; registered trademark of Unilever HPC-NA). The work-
ing hypothesis is depicted in Figure 15.8, which shows a TM film with a single oriented
layer of purple membrane and an ML film with five oriented layers of purple membranes
(in an actual experiment, it ranges from eight to ten layers). The
B
1
component, being
B
1
B
2
FIGURE 15.7
Schematic diagram showing the decomposition of the composite signal (
B
1
B
2
) when the access impedance is not zero. Each component decays in
two exponential terms as dictated by the equivalent circuit and the zero
time-integral condition. Note that inhibition of the
B
2
component results in
a decrease of the negative peak but an increase of the positive peak. An
inhibition of
B
2
alone could be misinterpreted as a concurrent enhancement
of
B
1
. (From Hong, F. T., Montal, M. (1979). Bacteriorhodopsin in model
membranes: a new component of the displacement photocurrent in the
microsecond time scale.
Biophys. J.
25:465-472.)
B
1
+
B
2
