Electrochemical Analysis of Ion Channels and Transporters in Pore-Suspending Membranes - Applications of Electrochemistry and Nanotechnology in Biology and Medicine - page 318

while that of the extracellular surface is calculated to be only -0.08
C m -2 at neutral pH. 100 This difference in surface charge density
would imply a preferential attachment of the cytoplasmic surface
to the bilayer, while the extracellular side faces the cis compart-
ment, which would in principle result in a proton transport direc-
tion from the nano-BLM towards the solution ( cis compartment).
Since the proton transport is observed in the opposite direction, at
least part of the purple membrane fragments are oriented in the
other direction and the current is dominated by this purple mem-
brane fraction. It is also well conceivable that not the surface
charge density but the different morphologies of the extracellular
and cytoplasmic side of the PM leads to a preferential adsorption,
with the extracellular side facing the nano-BLM.
To further prove that the observed proton currents are a result
of bR activity, an action spectrum was measured by monitoring the
initial current density using a series of narrow-band filters ( Fig. 18
B ). Compared to the absorption spectrum of purple membranes in
solution, the action spectrum obtained from purple membranes ad-
sorbed on nano-BLMs is shifted from 568 nm by about 20 nm to
548 nm. This observation is in accordance with previously pub-
lished results, where purple membranes were attached to classical
BLMs. 93 The authors monitored an action spectrum that was blue
shifted by about 15 nm compared to the absorption spectrum of
purple membranes in solution. Furthermore, we were able to iden-
tify the light dependence of the maximum current densities after
switching the light on and off, respectively ( Fig. 19 ). The pump
current I p,0 is defined as the current at V p = 0 and, therefore, I max
depends on the light intensity L according to Eq. (26):

L

I I

s

(26)

p,0 p,0

LL

1
2

where p, I is the saturation current of I p,0 for L Æ f and L 1/2 is the
half-saturation intensity. We observed that the activity of bR,
measured as the maximum current density under illumination as a
function of the relative light intensity, follows the theoretically
predicted course. Whilst J max,on is strongly dependent on L , J max,off
changes only slightly with L .