Structural basis for substrate recognition mechanism of ER glucosidase II

The endoplasmic reticulum (ER) possesses a sophisticated quality control system to proofread newly synthesized proteins. A series of N-linked oligosaccharide intermediates attached on the nascent proteins serves as specific tags for the quality control system. In this system, glucosidase II is involved in trimming of non-reducing terminal glucose residue of N-glycan intermediates. Glucosidase II consists of approximately 110 kDa catalytic α subunit (GIIα) and 60 kDa non-catalytic regulatory β subunit (GIIβ). It has been shown that GIIα alone can hydrolyze a small α-glycosidase model substrate (pNP-glucose), while it cannot catalyze deglucosylation of the N-linked oligosaccharide substrates unless it makes a complex with GIIβ. In this study, we determined the first crystal structure of GIIα in the absence and presence of its inhibitor 1-deoxynojirimycin at 1.6-Å resolution. The crystal structure revealed that GIIα has a characteristic segment at the N-terminus as compared with the cognate glycoside hydrolases (GH31). Interestingly, the N-terminal segment was accommodated on the substrate-binding pocket. Based on these results, we suggest that the N-terminal segment of GIIα undergoes structural rearrangement through interaction with GIIβ, thereby promoting the substrate-binding capacity for the N-linked oligosaccharide substrates.

Structural Basis for Emergence of Superionic Conductivity by an Ion Exchange

Various superionic conductors have been examined in terms of the application to electrolytes for solid fuel cells [1]. Recently we demonstrated by impedance measurements that a simple two-step chemical reaction transformed an electronic conductor Na_xCoO₂ into a superionic one. In the present study, we performed in situ synchrotron X-ray diffraction experiments to investigate a structural mechanism for the superionic conductivity driven by the chemical treatment of the layered oxide Na_xCoO₂. We developed a temperature- and humidity-controllable capillary cell under hydrogen and helium gas flow to install in the Debye-Scherrer camera at BL44B2 of SPring-8. This cell allows us to explore a structural transformation process by reduction and humidification treatments. Structural identifications and refinements with in situ diffraction data proved that Co vacancies formed by a CoO separation suppressed the electronic conductivity. Meanwhile it turned out from charge estimation in the Na layers that the superionic conductor transition originated from an ion exchange of H₃O⁺ for Na⁺, which was confirmed by Raman spectroscopy measurements. In addition, charge densities clearly visualized the H₃O⁺ ions disordering around the Na original sites, suggesting that the H₃O⁺ behave as a carrier source. Finally it was found from electrostatic potentials that the disordering H₃O⁺ sites were coupled through shallow potential barriers to trace a honeycomb-like ion pathway. In the presentation, I will discuss what a carrier is for the superionic-conductive phase from different viewpoints such as activation energies, concentration cell tests, and molecular dynamics simulations using the experimental structure information.

SAD experiment using the wavelength of 2.7 Å or longer

Single-wavelength anomalous dispersion (SAD) experiment with light atoms as anomalous scatterers has been carried out using longer wavelengths up to 2.3 Å. We have been developing a synchrotron beamline dedicated to the SAD experiments where wavelengths longer than 2.7 Å are available to enhance weak anomalous signals. Larger background noise due to the longer wavelength, which is one of the major problems in the experiment, is reduced by introducing a standing helium chamber surrounding both the whole diffractometer and the X-ray detector. The system allows to perform experiments with normal and long waveldngths under the same environment. Helium cold stream is fed into the chamber at the sample position and reused after removing contaminants to keep the temperature of the stream at 30 K or below economically. Capillary-top-mount method [1] was improved to further reduce the background noise and to accommodate with smaller or needle-shape crystals. Several results on de-novo structural solutions with sulfur-SAD phasing will be reported in addition to the current performance of the beamline and its future plan.

In situ synchrotron powder diffraction for an ionic conductor transition

In situ synchrotron X-ray powder diffraction can be one of the most powerful probes to investigate the structure evolution by a chemical reaction thanks to simultaneity of data collection. It is not, however, with ease to produce a homogeneous chemical reaction in the limited spaces, which is essential to see an atomic-scale structure evolution. We have developed an in situ capillary cell for both high-temperature H₂ reduction and precise humidity control at the SPring-8 BL44B2. We successfully applied this in situ system to an electronic conductor LaSr₃Fe₃O₁₀, which is transformed into an ionic conductor by the two-step chemical treatments [1]. LaSr₃Fe₃O₁₀ has a triple-layer structure with a FeO₆ octahedral unit. One triple layer is bonded with another layer through van der Waals interaction. Structure refinements with in situ synchrotron powder diffraction data revealed that the H₂ reduction at 613 K produced in-plane oxygen vacancies, which resulted in suppression of the interlayer interaction. We found from charge density studies and Raman spectroscopy measurements that the following humidification intercalated H₂O and OH^- into the interlayer and intralayer, respectively. That means that H₂O plays a role for suppression of three-dimensional electronic conductivity, stabilizing the intercalation structure. On the other hand, the OH^- ions behave as carriers for ionic conductivity, maintaining the charge neutrality in the intralayer. Finally we determined the composition of the ionic conductor to be LaSr₃Fe₃O_8.0 (OH) _1.2·2H₂O, which indicates a transformation of LaSr₃Fe₃O₁₀ into an OH^- ionic conductor. In the presentation, I will discuss the OH^- ionic conduction channel based on electrostatic potentials obtained from charge densities.

Orbital Order and Structural Phase Transitions in Vanadium Spinel FeV₂O₄

Orbital degrees of freedom plays an important role in condensed matter physics because it is strongly related with phase transitions and induces the fascinating physical properties. A spinel oxide FeV₂O₄ is one of the peculiar examples because this compound has double orbital degrees of freedom at both Fe²⁺ and V³⁺ ions. Furthermore, this material represents exotic physical properties [1,2], i.e.; multiferroic, large magnetostriction, and successive structural transitions with decreasing temperature: cubic - tetragonal (c < a: tetraHT, 138K) - orthorhombic (orthoHT, 108 K) - tetragonal (c > a: tetraLT, 68 K). However, the origin of structural transitions and physical properties is controversial until now. In order to clarify the origin, we have performed synchrotron x-ray diffraction experiments at low temperatures at beamline BL02B2 (for the powder samples) in SPring-8 and BL-4C (for the single crystal) of the Photon Factory, KEK. Furthermore, we have carried out the magnetization and the specific heat measurements using polycrystalline samples and single crystal of FeV₂O₄. We have firstly found another orthorhombic phase (orthoLT) below 30 K in the polycrystalline sample of FeV₂O₄, shown in figure 1. The Rietveld analysis was performed, and the overall qualities of fittings were fairly good. In order to investigate the details of the orbital state of Fe²⁺ and V³⁺ in FeV₂O₄, we have performed the normal mode analysis, which is based on static displacements of the tetrahedron of FeO₄ and octahedron of VO₆. In the orthoLT phase, we found the orbital order of Fe²⁺ ions, which is mixture of 3z²-r² and y²-z² orbitals, without change of orbital order of V³⁺ ions. This result indicates that the origin of the orthoLT phase is derived from the competition between cooperative Jahn-Teller effect and relativistic spin-orbit coupling of Fe²⁺ ions. We also discuss the origins of the other phase transitions considering the orbital state of V³⁺ and Fe²⁺ ions, and then the orbital dilution effect, where the structural and magnetic properties are investigated by using powder samples substituted for Fe²⁺ and V³⁺ ions by other ions (Mn²⁺ and Fe³⁺) with no orbital degrees of freedom.