578
Bacteriorhodopsin, Molecular Biology of
4
H
+
H
+
Asp96
Asp85
Arg82
Glu204
Glu194
Retinal
3
1
5
2
Fig. 1
Three-dimensional structure of
bacteriorhodopsin showing the seven
transmembrane helices, the retinal, and
functionally important residues as
marked. The arrows with numbers
represent proton transfer events in their
sequence. (Reproduced with permission
from Luecke et al. (1999)
Science
286
,
255–260.)
that becomes the acceptor of the Schiff
base proton. The pathway for the trans-
ported proton is a chain that leads from
Asp85 via Arg82, Glu204, and Glu194,
with seven intervening water molecules,
to the extracellular surface. In contrast,
the cytoplasmic half-channel contains hy-
drophobic residues between the retinal and
Asp96, the proton donor in the reprotona-
tion of the Schiff base. The side chain of
Thr46 forms a hydrogen bond with the
carboxyl group of Asp96, raising its p
K
a
high enough to keep it protonated. This
region is also connected to the retinal, but
through a chain of covalent bonds and
hydrogen bonds, with the participation of
part of helix G that forms a
π
–bulge near
Lys216, two water molecules, and Trp182
that contacts the retinal at the 13-methyl
group. The participants of the extracellular
and cytoplasmic chains are the candidates
for linking the events at the retinal Schiff
base Frst to proton release and then to
uptake at the two membrane surfaces.
3
Chromophore and the Photocycle
The chromophore with all-trans, 15-anti
retinal has a broad absorption band with a
maximum at 568 nm. This is considerably
red-shifted from that of retinal (380 nm) or
a retinal analogue with a protonated Schiff
base (440 nm). The ‘‘opsin shift’’ origi-
nates from the only partial compensation
of the positively charged retinal Schiff base
by the counterion complex, and the distor-
tion of the polyene chain of the retinal in
its binding site. The all-trans chromophore
exists in thermal equilibrium with the 13-
cis,15-syn conFguration, which absorbs
at 555 nm. Sustained illumination con-
verts the retinal to 100% all-trans, 15-anti
(‘‘light-adaptation’’), and it is the photore-
action of this isomer that is normally active
in transport.
The photochemical cycle of the all-trans
chromophore is described by the interme-
d
iatestatesJ
,K
,L
,M
,N
,andOandthe
ir
substates and the sequence of their in-
terconversions (±ig. 2). Each intermediate
is characterized by a distinct absorption
maximum in the visible and numerous
vibrational bands from the retinal in the
infrared and Raman, as well as from the
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