Highly Selective Separation of Rhodopsin from Bovine Rod Outer Segment Membranes Using Combination of Divalent Cation and Alkyl(thio)glucoside

The micellization process of bovine rod outer segment (ROS) membranes is investigated utilizing a series of neutral detergents. It is found that when alkyl(thio)glucosides with an appropriate hydrophilic–lipophilic balance (e.g. octylthioglucoside) are used in combination with a divalent cation, rhodopsin is selectively extracted from ROS membranes at a specific detergent-to-membrane ratio. This allows remarkable purification of rhodopsin by a single-step solubilization, because the residual membranes are heavily aggregated in the presence of divalent cation and are therefore easily sedimented by low-speed centrifugation. The absorption spectrum of the supernatant reproducibly exhibits an A₂₈₀/A₅₀₀ value of 1.6, an excellent value that could rarely be obtained by chromatographic purification. The degree of purification also depends on the type of divalent cation included in the solubilization solution; specific binding of IIB-series cations (Zn²⁺ and Cd²⁺) to ROS membranes is suggested to play an important role in the solubilization process. The present result represents a unique example of selective solubilization of a specific membrane protein from highly aggregated membranes.     

Direct observations of freeze-etching processes of ice-embedded biomembranes by atomic force microscopy

We have fabricated a cryogenic atomic force microscope that is designed for structural investigation of freeze-fractured biological specimens. The apparatus is operated in liquid nitrogen gas at atmospheric pressure. Freeze-fracturing, freeze-etching and subsequent imaging are carried out in the same chamber, so that the surface topography of a fractured plane is easily visualized without ice contamination. A controlled superficial sublimation of volatile molecules allows us to obtain three-dimensional views of ultrastructures of biological membranes.     

IR laser manipulation of cis<-->trans isomerization of 2-naphthol and its hydrogen-bonded clusters

The cis<-->trans isomerization reaction has been carried out for 2-naphthol and its hydrogen (H) bonded clusters by infrared (IR) laser in the electronic excited state (S1) in supersonic jets. A specific isomer in the jet was pumped to the X-H stretching vibration in the S1 state, where X refers to C, O, or N atom, by using a stepwise UV-IR excitation, and the dispersed emission spectra of the excited species or generated fragments were observed. It was found that the isomerization occurs only in the H-bonded clusters but a bare molecule does not exhibit the isomerization in the examined energy region of Ev< or =3610 cm(-1), indicating a reduction of the isomerization barrier height upon the H bonding. The relative yield of the isomerization was observed as a function of internal energy. The isomerization yield was found to be very high at the low IR frequency excitation, and was rapidly reduced with the IR frequency due to the competition of the dissociation of the H bond within the isomer. Density-functional theory (DFT) and time-dependent DFT calculations were performed for estimating the barrier height of the isomerization for bare 2-naphthol and its cluster for electronic ground and excited states. The calculation showed that the isomerization barrier height is highly dependent on the electronic states. However, the reduction of the height upon the hydrogen bonding was not suggested at the level of our calculation.     

FORMATION AND DECAY KINETICS OF BACTERIORHODOPSIN'S N INTERMEDIATE: SOFTENING THE PROTEIN CONFORMATION WITH ALCOHOLS AFFECTS INTRA-PIROTEIN PROTON TRANSFER AND RETINAL ISIOMERIZATION

Abstract— The trans photocycle of bacteriorhodopsin was investigated in the presence of organic solvents with a hydrogen-bonding group; i.e . methanol, ethanol, 1-propanol and so on. These alcohols scarcely or only slightly affected the L→M and O₅₇₀ transitions, but they perturbed the M→N and N→O transitions greatly. The rate of the M→N transition increased linearly with increasing alcohol concentration and, at maximal alcohol concentrations under which the native protein conformation was retained, the M→N transition was accelerated by a factor of ∼5. This alcohol effect was reversible. It is suggested that a long-distance proton transfer involved in the M→N transition (Asp.96→retinal) becomes easier when the protein conformation is softened bv partially breaking hydrogen-bonding networks in the protein. Another significant effect of alochol is inhibition of the N→O transition at weakly acidic pH, which was slowed down maximally by a factor of ˜10. This alcohol effect was less significant at alkaline pH, where reprotonation of Asp-96 from the cytoplasmic membrane surface is a rate-limiting reaction. It is suggested that, at acidic pH, thi: cis-to-trans isomerization involved in the N→O transition is a rate-limiting reaction and that this step is inhibited in the presence of a high concentration of alcohols.     

Photoluminescence of exciton and biexciton in GaN nanocolumns

We measured photoluminescence (PL) from excitons and biexcitons in GaN nanocolumns at low temperature and found that the PL spectra of excitons depend on the nanocolumn diameter. Taking into account the polaritonic effect of the excitons, calculation of the PL spectra revealed that the dependence on diameters causes a difference of PL intensity from side surfaces of the nanocolumns. At high excitation intensities, we also observed biexciton emissions and found that the biexciton binding energies are higher than those in bulk samples. Although the mechanism for the increase in the binding energy is not clear at present, we suppose that it arises from a spatial confinement effect due to the nanocolumn morphology.     

Transient differential reflectivity of ferromagnetic and paramagnetic phases in the bilayered manganite La1.24Sr1.76Mn2O7

Photoinduced effects in a single crystal of bilayered manganites, La_2−2xSr_1+2xMn₂O₇ (x=0.38), were investigated in a wide range of temperatures by pump-probe measurement at a photon energy of 1.6 eV. In a ferromagnetic metallic state, significant enhancement of positive rise in differential reflectivity with a slow relaxing time of 100 ps was observed just below T_C=127 K, indicating that the reflectivity change with the slow relaxation time constant is induced by laser heating. We have also observed an unconventional fast relaxing component that has a time constant of the order of 10 ps. This fast relaxing component, whose absolute value has an asymmetric peak at T_C, is presumably due to short-range correlation of Jahn-Teller distortion.     

Evaluation of bandgap energy and carrier density of InN nanocolumns

We have measured the optical properties of wurtzite InN nanocolumns and film by photoluminescence (PL) measurements at temperatures from 5 to 300 K and analyzed the PL spectra by fitting with the free-electron recombination bound (FERB) model. For the top-linked InN nanocolumns, we observed strong PL intensity compared to the InN film sample. The PL spectra were asymmetrical with low-energy tails and a red-shift of the PL peak energy position was observed with increasing temperature. However, for the separated InN nanocolumns, we observed weak PL intensity and symmetrical PL spectra. Analyzing the spectra shape of the top-linked InN nanocolumns at 5 K using the FERB model, we evaluated the intrinsic bandgap energy and carrier density of InN nanocolumns to be 0.69 eV and 2.5×10¹⁷ cm⁻³, respectively.     

Preparation of amphotropic liquid crystalline poly(alkoxycarbonylphenylene)s

Poly(alkoxycarbonylphenylene)s with long alkoxy groups were prepared by dehalogenative polycondensation of alkyl dichlorobenzoates with nickel bromide/zinc/triphenylphosphine catalyst. The poly(alkoxycarbonyl‐p‐phenylene)s and poly(alkyloxycarbonyl‐m‐phenylene)s are liquid crystalline, although the latter are composed wholly of kinked repeating units. While poly(hexyloxycarbonylphenylene)s exhibit only thermotropic liquid crystalline behavior, poly(dodecyloxycarbonylphenylene)s and poly(hexadecyloxycarbonylphenylene)s show amphotropism.