Superresolved image reconstruction in FZA lensless camera by color-channel synthesis.

The Fresnel-zone-aperture lensless camera using a fringe-scanning technique allows non-iterative well-conditioned image reconstruction; however, the spatial resolution is limited by the mathematical reconstruction model that ignores diffraction. To solve this resolution problem, we propose a novel image-reconstruction algorithm using the wave-optics-based design of the deconvolution filter and color-channel image synthesis. We verify a two-fold improvement of the effective angular resolution by conducting numerical simulations and optical experiments with a prototype.

Conductance of P2X4 purinergic receptor is determined by conformational equilibrium in the transmembrane region.

Ligand-gated ion channels are partially activated by their ligands, resulting in currents lower than the currents evoked by the physiological full agonists. In the case of P2X purinergic receptors, a cation-selective pore in the transmembrane region expands upon ATP binding to the extracellular ATP-binding site, and the currents evoked by α,ß-methylene ATP are lower than the currents evoked by ATP. However, the mechanism underlying the partial activation of the P2X receptors is unknown although the crystal structures of zebrafish P2X4 receptor in the apo and ATP-bound states are available. Here, we observed the NMR signals from M339 and M351, which were introduced in the transmembrane region, and the endogenous alanine and methionine residues of the zebrafish P2X4 purinergic receptor in the apo, ATP-bound, and α,ß-methylene ATP-bound states. Our NMR analyses revealed that, in the α,ß-methylene ATP-bound state, M339, M351, and the residues that connect the ATP-binding site and the transmembrane region, M325 and A330, exist in conformational equilibrium between closed and open conformations, with slower exchange rates than the chemical shift difference (<100 s(-1)), suggesting that the small population of the open conformation causes the partial activation in this state. Our NMR analyses also revealed that the transmembrane region adopts the open conformation in the state bound to the inhibitor trinitrophenyl-ATP, and thus the antagonism is due to the closure of ion pathways, except for the pore in the transmembrane region: i.e., the lateral cation access in the extracellular region.

Deuteration and selective labeling of alanine methyl groups of ß2-adrenergic receptor expressed in a baculovirus-insect cell expression system.

G protein-coupled receptors (GPCRs) exist in equilibrium between multiple conformations, and their populations and exchange rates determine their functions. However, analyses of the conformational dynamics of GPCRs in lipid bilayers are still challenging, because methods for observations of NMR signals of large proteins expressed in a baculovirus-insect cell expression system (BVES) are limited. Here, we report a method to incorporate methyl-13C1H3-labeled alanine with > 45% efficiency in highly deuterated proteins expressed in BVES. Application of the method to the NMR observations of ß2-adrenergic receptor in micelles and in nanodiscs revealed the ligand-induced conformational differences throughout the transmembrane region of the GPCR.

Efficient tiled calculation of over-10-gigapixel holograms using ray-wavefront conversion.

In the calculation of large-scale computer-generated holograms, an approach called "tiling," which divides the hologram plane into small rectangles, is often employed due to limitations on computational memory. However, the total amount of computational complexity severely increases with the number of divisions. In this paper, we propose an efficient method for calculating tiled large-scale holograms using ray-wavefront conversion. In experiments, the effectiveness of the proposed method was verified by comparing its calculation cost with that using the previous method. Additionally, a hologram of 128K × 128K pixels was calculated and fabricated by a laser-lithography system, and a high-quality 105 mm × 105 mm 3D image including complicated reflection and translucency was optically reconstructed.

Off-axis virtual-image display and camera by holographic mirror and blur compensation.

We propose off-axis virtual-image display and camera systems, which integrate a vertically-standing holographic off-axis mirror, blur-compensation optical systems, and digital imaging devices. In the system, the holographic mirror is used for an off-axis reflector, which realizes an upright and thin screen for virtual-image formation. By combining it with a display unit, an off-axis virtual-image display is realized, where the virtual image can be seen behind the upright holographic mirror. Simultaneously, by combining it with a camera unit, an off-axis camera is implemented, which realizes frontal shooting of objects by a camera placed at an off-axis position. Since both the off-axis display and the camera can be implemented by a single holographic mirror, it can be applied to a two-way visual-telecommunication system with a thin screen, which implements eye contact and the observer--image distance. A problem with the proposed system is image blur, which is caused by the chromatic dispersion of the holographic mirror. To solve this, we designed optical blur-compensation systems using a diffractive optical element and a diffuser or a lens. Experimental results verify the concept of the proposed systems with clarifying the effect of designed blur-compensation methods.

Non-diffracting linear-shift point-spread function by focus-multiplexed computer-generated hologram.

An Airy beam can be used to implement a non-diffracting self-bending point-spread function (PSF), which can be utilized for computational 3D imaging. However, the parabolic depth-dependent spot trajectory limits the range and resolution in rangefinding. In this Letter, we propose a novel pupil-phase-modulation method to realize a non-diffracting linear-shift PSF. For the modulation, we use a focus-multiplexed computer-generated hologram, which is calculated by multiplexing multiple lens-function holograms with 2D sweeping of the foci. With this method, the depth-dependent trajectory of the non-diffracting spot is straightened, which improves the range and resolution in rangefinding. The proposed method was verified by numerical simulations and optical experiments. The method can be applied to laser-based microscopy, time-of-flight rangefinding, and so on.

Fast method of calculating a photorealistic hologram based on orthographic ray-wavefront conversion.

A computer-generated hologram based on ray-wavefront conversion can reconstruct photorealistic three-dimensional (3D) images containing deep virtual objects and complicated physical phenomena; however, the required computational cost has been a problem that needs to be solved. In this Letter, we introduce the concept of an orthographic projection in the ray-wavefront conversion technique for reducing the computational cost without degrading the image quality. In the proposed method, plane waves with angular spectra of the object are obtained via orthographic ray sampling and Fourier transformation, and only the plane waves incident on the hologram plane are numerically propagated. We verified this accelerated computational method theoretically and experimentally, and demonstrated optical reconstruction of a deep 3D image in which the effects of occlusions, transmission, refraction, and reflection were faithfully reproduced.

Structure determination of a protein assembly by amino acid selective cross-saturation.

Amino acid selective cross-saturation (ASCS) method not only provides information about the interface of a protein assembly by the spin relaxation experiment, but also identifies the amino acid residues in the acceptor protein, which are located close to the selectively labeled amino acid residues in the donor protein. Here, a new method was developed to build a precise structural model of a protein assembly, which satisfies the experimental ASCS values, using simulated annealing computation. This method was applied to the ubiquitin-yeast ubiquitin hydrolase 1 (Ub-YUH1) complex to build a precise complex structure compatible with that determined by X-ray crystallography.

Amino acid selective cross-saturation method for identification of proximal residue pairs in a protein-protein complex.

We describe an NMR-based approach, the amino acid selective cross-saturation (ASCS) method, to identify the pairs of the interface residues of protein-protein complexes. ASCS uses a "cross-saturation (CS)-donor" protein, in which only one amino acid is selectively (1)H-labeled in a (2)H-background, and a "CS-acceptor" protein with uniform (2)H, (15)N labeling. Irradiation of the (1)H-labeled amino acid, which exists only in the donor, decreases the intensity of the (1)H- (15)N HSQC signals of the acceptor residues proximal to the (1)H-labeled CS-source residue(s) through the CS phenomenon. Given the three-dimensional structure of each protein in the complex, but not the complex structure, the combinatorial analysis of multiple ASCS results specify the CS-source residue(s), based on the spatial complementarity between the CS-source residues on the CS donor and the cross-saturated amide protons on the acceptor. NMR investigations of the labeling selectivity and efficiency in an E. coli host, which are critical for ASCS, revealed that Ala, Arg, His, Ile, Leu, Lys, Met, Phe, Pro, Trp, and Tyr are selectively labeled with a high (1)H/(2)H ratio. The observation of the ASCS was then confirmed using the known structure of the yeast ubiquitin (Ub) and yeast ubiquitin hydrolase 1 (YUH1). Conversely, reasonable candidates for the CS-source residues were suggested by the analysis of the ASCS results, with reference to the individual structures of YUH1 and Ub. The pairwise distance information between the CS-source residues and the cross-saturated amide groups obtained by ASCS will be useful for modeling protein-protein complexes.

Interaction Analysis of FABP4 Inhibitors by X-ray Crystallography and Fragment Molecular Orbital Analysis.

X-ray crystal structural determination of FABP4 in complex with four inhibitors revealed the complex binding modes, and the resulting observations led to improvement of the inhibitory potency of FABP4 inhibitors. However, the detailed structure-activity relationship (SAR) could not be explained from these structural observations. For a more detailed understanding of the interactions between FABP4 and inhibitors, fragment molecular orbital analyses were performed. These analyses revealed that the total interfragment interaction energies of FABP4 and each inhibitor correlated with the ranking of the K i value for the four inhibitors. Furthermore, interactions between each inhibitor and amino acid residues in FABP4 were identified. The oxygen atom of Lys58 in FABP4 was found to be very important for strong interactions with FABP4. These results might provide useful information for the development of novel potent FABP4 inhibitors.

Backbone resonance assignments for the ligand binding subunit of the histidine permease complex (HisJ) from Escherichia coli, under histidine-bound and unbound states.

HisJ is a histidine binding subunit of the histidine permease, which exists in the outer membrane of Gram-negative bacteria. In order to incorporate the periplasmic histidine into the cell, HisJ captures histidine in the periplasm, and transfers the histidine to the transmembrane complex of histidine permease that is an ABC transporter. We established the backbone resonance assignments of (1)H/(13)C/(15)N-labeled HisJ from Escherichia coli, in the histidine-bound and unbound states.