11 research outputs found
Electron Conduction and Photocurrent Generation of a Light-Harvesting/Reaction Center Core Complex in Lipid Membrane Environments
To reveal the structure–function
relationship of membrane
proteins, a membrane environment is often used to establish a suitable
platform for assembly, functioning, and measurements. The control
of the orientation of membrane proteins is the main challenge. In
this study, the electron conductivity and photocurrent of a light-harvesting/reaction
center core complex (LH1-RC) embedded in a lipid membrane were measured
using conductive atomic force microscopy (C-AFM) and photoelectrochemical
analysis. AFM topographs showed that LH1-RC molecules were well-orientated,
with their H-subunits toward the membrane surface. Rectified conductivity
was observed in LH1-RC under precise control of the applied force
on the probe electrode (<600 pN). LH1-RC embedded in a membrane
generated photocurrent upon irradiation when assembled on an electrode.
The observed action spectrum was consistent with the absorption spectrum
of LH1-RC. The control of the orientation of LH1-RC by lipid membranes
provided well-defined conductivity and photocurrent
Gating-Associated Clustering–Dispersion Dynamics of the KcsA Potassium Channel in a Lipid Membrane
The KcsA potassium channel is a prototypical channel of bacterial
origin, and the mechanism underlying the pH-dependent gating has been
studied extensively. With the high-resolution atomic force microscopy
(AFM), we have resolved functional open and closed gates of the KcsA
channel under the membrane-embedded condition. Here we surprisingly
found that the pH-dependent gating of the KcsA channels was associated
with clustering–dispersion dynamics. At neutral pH, the resting,
closed channels were coalesced, forming nanoclusters. At acidic pH,
the open-gated channels were dispersed as singly isolated channels.
Time-lapse AFM revealed reversible clustering–dispersion transitions
upon pH changes. At acidic equilibrium, a small fraction of the channels
was nanoclustered, in which the gate was apparently closed. Thus,
it is suggested that opening of the gate and the dispersion are tightly
linked. The interplay between the intramolecular conformational change
and the supramolecular clustering–dispersion dynamics provides
insights into understanding of unprecedented functional cooperativity
of channels
Gating-Associated Clustering–Dispersion Dynamics of the KcsA Potassium Channel in a Lipid Membrane
The KcsA potassium channel is a prototypical channel of bacterial
origin, and the mechanism underlying the pH-dependent gating has been
studied extensively. With the high-resolution atomic force microscopy
(AFM), we have resolved functional open and closed gates of the KcsA
channel under the membrane-embedded condition. Here we surprisingly
found that the pH-dependent gating of the KcsA channels was associated
with clustering–dispersion dynamics. At neutral pH, the resting,
closed channels were coalesced, forming nanoclusters. At acidic pH,
the open-gated channels were dispersed as singly isolated channels.
Time-lapse AFM revealed reversible clustering–dispersion transitions
upon pH changes. At acidic equilibrium, a small fraction of the channels
was nanoclustered, in which the gate was apparently closed. Thus,
it is suggested that opening of the gate and the dispersion are tightly
linked. The interplay between the intramolecular conformational change
and the supramolecular clustering–dispersion dynamics provides
insights into understanding of unprecedented functional cooperativity
of channels
Lipid-Controlled Stabilization of Charge-Separated States (P<sup>+</sup>Q<sub>B</sub><sup>–</sup>) and Photocurrent Generation Activity of a Light-Harvesting–Reaction Center Core Complex (LH1-RC) from <i>Rhodopseudomonas palustris</i>
The
photosynthetic light-harvesting–reaction center core
complex (LH1-RC) is a natural excitonic and photovoltaic device embedded
in a lipid membrane. In order to apply LH1-RCs as a biohybrid energy-producing
material, some important issues must be addressed, including how to
make LH1-RCs function as efficiently as possible. In addition, they
should be characterized to evaluate how many active LH1-RCs efficiently
work in artificial systems. We report here that an anionic phospholipid,
phosphatidylglycerol (PG), stabilizes the charge-separated state (a
photooxidized electron donor and reduced quinone pair, P<sup>+</sup>Q<sub>B</sub><sup>–</sup>) of LH1-RC (from <i>Rhodopseudomonas
palustris</i>) and enhances its activity in photocurrent generation.
Steady-state fluorometric analysis demonstrated that PG enhances the
formation of the P<sup>+</sup>Q<sub>B</sub><sup>–</sup> state
at lower irradiances. The photocurrent generation activity was analyzed
via Michaelis–Menten kinetics, revealing that 38% of LH1-RCs
reconstituted into the PG membrane generated photocurrent at a turnover
frequency of 46 s<sup>–1</sup>. PG molecules, which interact
with LH1-RC in vivo, play the role of an active effector component
for LH1-RC to enhance its function in the biohybrid system
Oxygen-Evolving Porous Glass Plates Containing the Photosynthetic Photosystem II Pigment–Protein Complex
The
development of artificial photosynthesis has focused on the
efficient coupling of reaction at photoanode and cathode, wherein
the production of hydrogen (or energy carriers) is coupled to the
electrons derived from water-splitting reactions. The natural photosystem
II (PSII) complex splits water efficiently using light energy. The
PSII complex is a large pigment–protein complex (20 nm in diameter)
containing a manganese cluster. A new photoanodic device was constructed
incorporating stable PSII purified from a cyanobacterium Thermosynechococcus vulcanus through immobilization
within 20 or 50 nm nanopores contained in porous glass plates (PGPs).
PSII in the nanopores retained its native structure and high photoinduced
water splitting activity. The photocatalytic rate (turnover frequency)
of PSII in PGP was enhanced 11-fold compared to that in solution,
yielding a rate of 50–300 mol e<sup>–</sup>/(mol PSII·s)
with 2,6-dichloroindophenol (DCIP) as an electron acceptor. The PGP
system realized high local concentrations of PSII and DCIP to enhance
the collisional reactions in nanotubes with low disturbance of light
penetration. The system allows direct visualization/determination
of the reaction inside the nanotubes, which contributes to optimize
the local reaction condition. The PSII/PGP device will substantively
contribute to the construction of artificial photosynthesis using
water as the ultimate electron source
Influence of Phospholipid Composition on Self-Assembly and Energy-Transfer Efficiency in Networks of Light-Harvesting 2 Complexes
In
the photosynthetic membrane of purple bacteria networks of light-harvesting
2 (LH2) complexes capture the sunlight and transfer the excitation
energy. In order to investigate the mutual relationship between the
supramolecular organization of the pigment–protein complexes
and their biological function, the LH2 complexes were reconstituted
into three types of phospholipid membranes, consisting of l-α-phosphatidylglycerol (PG), l-α-phosphatidylcholine
(PC), and l-α-phosphatidylethanolamine (PE)/PG/cardiolipin
(CL). Atomic force microscopy (AFM) revealed that the type of phospholipids
had a crucial influence on the clustering tendency of the LH2 complexes
increased from PG over PC to PE/PG/CL, where the LH2 complexes formed
large, densely packed clusters. Time-resolved spectroscopy uncovered
a strong quenching of the LH2 fluorescence that is ascribed to singlet–singlet
and singlet–triplet annihilation by an efficient energy transfer
between the LH2 complexes in the artificial membrane systems. Quantitative
analysis reveals that the intercomplex energy transfer efficiency
varies strongly as a function of the morphology of the nanostructure,
namely in the order PE/PG/CL > PC > PG, which is in line with
the
clustering tendency of LH2 observed by AFM. These results suggest
a strong influence of the phospholipids on the self-assembly of LH2
complexes into networks and concomitantly on the intercomplex energy
transfer efficiency
Light-Driven Hydrogen Production by Hydrogenases and a Ru-Complex inside a Nanoporous Glass Plate under Aerobic External Conditions
Hydrogenases
are powerful catalysts for light-driven H<sub>2</sub> production using
a combination of photosensitizers. However, except
oxygen-tolerant hydrogenases, they are immediately deactivated under
aerobic conditions. We report a light-driven H<sub>2</sub> evolution
system that works stably even under aerobic conditions. A [NiFe]-hydrogenase
from <i>Desulfovibrio vulgaris</i> Miyazaki F was immobilized
inside nanoporous glass plates (PGPs) with a pore diameter of 50 nm
together with a ruthenium complex and methyl viologen as a photosensitizer
and an electron mediator, respectively. After immersion of PGP into
the medium containing the catalytic components, an anaerobic environment
automatically established inside the nanopores even under aerobic
external conditions upon irradiation with solar-simulated light; this
system constantly evolved H<sub>2</sub> with an efficiency of 3.7
μmol H<sub>2</sub> m<sup>–2</sup> s<sup>–1</sup>. The PGP system proposed in this work represents a promising first
step toward the development of an O<sub>2</sub>-tolerant solar energy
conversion system
Extension of Light-Harvesting Ability of Photosynthetic Light-Harvesting Complex 2 (LH2) through Ultrafast Energy Transfer from Covalently Attached Artificial Chromophores
Introducing appropriate artificial
components into natural biological
systems could enrich the original functionality. To expand the available
wavelength range of photosynthetic bacterial light-harvesting complex
2 (LH2 from Rhodopseudomonas acidophila 10050), artificial fluorescent dye (Alexa Fluor 647: A647) was covalently
attached to N- and C-terminal Lys residues in LH2 α-polypeptides
with a molar ratio of A647/LH2 ≃ 9/1. Fluorescence and transient
absorption spectroscopies revealed that intracomplex energy transfer
from A647 to intrinsic chromophores of LH2 (B850) occurs in a multiexponential
manner, with time constants varying from 440 fs to 23 ps through direct
and B800-mediated indirect pathways. Kinetic analyses suggested that
B800 chromophores mediate faster energy transfer, and the mechanism
was interpretable in terms of Förster theory. This study demonstrates
that a simple attachment of external chromophores with a flexible
linkage can enhance the light harvesting activity of LH2 without affecting
inherent functions of energy transfer, and can achieve energy transfer
in the subpicosecond range. Addition of external chromophores, thus,
represents a useful methodology for construction of advanced hybrid
light-harvesting systems that afford solar energy in the broad spectrum
Molecular Assembly of Zinc Chlorophyll Derivatives by Using Recombinant Light-Harvesting Polypeptides with His-tag and Immobilization on a Gold Electrode
LH1-α
and -β polypeptides, which make up the light-harvesting
1 (LH1) complex of purple photosynthetic bacteria, along with bacteriochlorophylls,
have unique binding properties even for various porphyrin analogs.
Herein, we used the porphyrin analogs, Zn-Chlorin and the Zn-Chlorin
dimer, and examined their binding behaviors to the LH1-α variant,
which has a His-tag at the C-terminus (MBP-rubα-YH). Zn-Chlorin
and the Zn-Chlorin dimer could bind to MBP-rubα-YH and form
a subunit-type assembly, similar to that from the native LH1 complex.
These complexes could be immobilized onto Ni-nitrilotriacetic acid-modified
Au electrodes, and the cathodic photocurrent was successfully observed
by photoirradiation. Since Zn-Chlorins in this complex are too far
for direct electron transfer from the electrode, a contribution of
polypeptide backbone for efficient electron transfer was implied.
These findings not only show interesting properties of LH1-α
polypeptides but also suggest a clue to construct artificial photosynthesis
systems using these peptide materials
Immobilization and Photocurrent Activity of a Light-Harvesting Antenna Complex II, LHCII, Isolated from a Plant on Electrodes
A light-harvesting (LH) antenna complex II, LHCII, isolated
from
spinach was immobilized onto an indium tin oxide (ITO) electrode with
dot patterning of 3-aminopropyltriethoxysilane (APS) by utilizing
electrostatic interactions between the cationic surface of the electrode
and the anionic surface of stromal side of the LHCII polypeptide.
Interestingly, the illumination of LHCII assembled onto the ITO electrode
produced a photocurrent response that depends on the wavelength of
the excitation light. Further, LHCII was immobilized onto a TiO<sub>2</sub> nanostructured film to extend for the development of a dye-sensitized
biosolar cell system. The photocurrent measured in the iodide/tri-iodide
redox system of an ionic liquid based electrolyte on the TiO<sub>2</sub> system showed remarkable enhancement of the conversion efficiency,
as compared to that on the ITO electrode