Bacteriorhodopsin, Molecular Biology of
Bacteriorhodopsin is a small (26 kDa) in-
tegral membrane protein, the prototype of
seven-helical G-protein-linked receptors.
It is found in extended two-dimensional
hexagonal arrays in the cytoplasmic mem-
brane of halobacteria, the ‘‘purple mem-
brane’’ patches. Upon illumination, this
retinal-containing protein transports pro-
tons against a transmembrane gradient.
Photoisomerization of the retinal from all-
trans to 13-cis,15-anti sets off a sequence
of thermal reactions, the ‘‘photocycle,’’ in
which changes of the retinal cause a cy-
cle of structural changes in the protein,
and these result in vectorial proton trans-
fers between donor and acceptor groups.
Together, the internal transfers add up to
full translocation of a proton from the
cytoplasmic to the extracellular side of
the protein and thus generate a trans-
membrane electrochemical gradient for
protons. This light-driven gradient is uti-
lized for chemiosmotic coupling, in the
same way as in other membranes in which
it is generated by redox reactions, to the
synthesis of ATP, the uptake of K
nutrients (amino acids), and the transport
of Na
out of the cells.
Overall Structure of the Protein
Bacteriorhodopsin forms trimers, which
assemble in the two-dimensional hexago-
nal lattice of the purple membrane. This
specialized membrane contains only bac-
teriorhodopsin and speciFc lipids (about
10 lipids/protein), and its regular crys-
talline lattice made it possible to determine
the structure of the protein by cryoelectron
microscopy of single sheets, ultimately to
A resolution. Three-dimensional crystals
can be grown from detergent-solubilized
bacteriorhodopsin in cubic lipid phase,
and from X-ray diffraction the structure of
the protein has been described at increas-
ingly better resolutions, most recently to
A. ±rom these crystallographic stud-
ies, the protein is known to consist of
seven transmembrane helices with short
interhelical loops and short N and C ter-
mini. Three of the helices, B, C, and D,
are normal to the plane of the membrane
and four, A, E, ±, and G, are inclined at
various but small angles to the perpendicu-
lar. The single retinal per protein molecule
is bound to the
-amino group of Lys216
forming a protonated Schiff base near the
lies at a small angle from the membrane
plane (±ig. 1).
The Schiff base divides the seven-helical
bundle into extracellular and cytoplasmic
halves. The trajectory of the transported
proton is through the two ‘‘half-channels’’
formed by these regions. IdentiFcation of
the residues and bound water that par-
ticipate in these half-channels has been
the objective of much work in the last
few decades. Their locations and how
they interact are known from crystallogra-
phy, infrared spectroscopy, and solid-state
nuclear magnetic resonance (NMR). The
extracellular half-channel contains numer-
ous polar and hydrogen-bonding residues
and a three-dimensional network of tightly
shown that the polar side chains play roles
in the release of protons to the extracellular
surface. The anionic Asp85 and Asp212 are
hydrogen-bonded to the centrally located
water402 that receives a hydrogen bond
from the protonated retinal Schiff base.
The Asp85/Asp212/water402 complex is
the counterion to the charge of the Schiff
base, but in the photocycle it is Asp85
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