研究者詳細 - 内橋　貴之

論文 - 内橋　貴之

分割表示 237 件中 1 - 20 件目／全件表示 >>

Characterization of high affinity IgM and IgG monoclonal antibodies against norovirus variants GII.4 and GII.17 査読有り Open Access

Tagawa J., Yanaka S., Kato Y., Masuda A., Lee J.M., Senoo A., Oyama K., Uchihashi T., Nishida M., Kusakabe T., Caaveiro J.M.M.

Protein Science 35 巻 ( 4 ) 頁： e70522 2026年4月

記述言語：英語出版者・発行元：Protein Science

Human noroviruses are a leading cause of acute gastroenteritis worldwide, yet the molecular principles governing antibody recognition of their highly repetitive capsid remain poorly understood. Here, we immunized mice with virus-like particles (VLPs) from the pandemic GII.4 strain and the emergent GII.17 strain, generating monoclonal IgM and IgG antibodies via hybridoma technology. High-speed atomic force microscopy visualized IgM antibodies scanning and engaging multiple protruding (P) domains on intact VLPs. Surface plasmon resonance (SPR) analyses of engineered antibodies with identical Fab sequences but different valencies revealed that, unlike monovalent IgG, multivalent IgM exhibits dramatic affinity gains—up to 100-fold—as antigen density increases. This avidity-driven enhancement arises from the dense, repetitive P-domain architecture of the norovirus capsid, enabling IgM to achieve high functional affinity despite modest intrinsic Fab binding. Our findings define how antibody valency and epitope organization cooperate to boost viral recognition, offering a mechanistic framework for designing next-generation vaccines and antiviral antibodies that harness multivalent engagement.

DOI： 10.1002/pro.70522

Key Role of Pro230 in the Hinge Region on the Architecture and Function of IgG1 査読有り Open Access

Koseki, Y; Yamaguchi, Y; Aoyama, M; Hiraka, K; Tada, M; Kodama, A; Senoo, A; Ishii-Watabe, A; Uchihashi, T; Murata, K; Uchiyama, S; Kato, K; Yanaka, S; Caaveiro, JMM

JOURNAL OF MEDICINAL CHEMISTRY 69 巻 ( 4 ) 頁： 3902 - 3914 2026年2月

記述言語：英語出版者・発行元：Journal of Medicinal Chemistry

Immunoglobulin G (IgG) is a multifunctional glycoprotein essential for immune defense and widely used as a therapeutic due to its antigen specificity and effector functions. However, the inherent flexibility of its hinge region complicates structural characterization and obscures the molecular basis of its mechanism of action. To clarify the hinge’s role, we performed systematic amino acid substitutions. Notably, deletion of Pro230 led to the formation of a half-IgG1 species lacking inter-heavy chain interactions. Structural analysis using nuclear magnetic resonance (NMR), negative-stain EM, and disulfide bond quantification by LC-MS/MS peptide mapping revealed the mechanism underlying half-IgG1 generation. To enable this, we developed a new stable-isotope labeling method for NMR. Functional assays with FcγR-expressing reporter cells demonstrated that half-IgG1 retained selective FcγRI-mediated activity. These findings provide new insights into higher-order IgG structure and Fcγ receptor-dependent immune activation, offering a basis for designing next-generation antibody therapeutics.

DOI： 10.1021/acs.jmedchem.5c02419

Real-Time Sub-Molecular Visualization of DNA Folding Using a Hybrid High-Speed AFM and Optical Tweezers System 査読有り

Umeda, K; Yamanaka, S; Imamura, M; Nagae, F; Fukuda, S; Watanabe, H; Uchihashi, T; Takada, S; Ando, T

JOURNAL OF PHYSICAL CHEMISTRY LETTERS 17 巻 ( 4 ) 頁： 1182 - 1191 2026年1月

記述言語：英語掲載種別：研究論文（学術雑誌）出版者・発行元：Journal of Physical Chemistry Letters

Optical tweezers have contributed to elucidating the folding mechanisms associated with biomolecules. By combining them with fluorescence imaging techniques, imaging can also be performed while measuring or applying forces that couple to biochemical reactions; however, they cannot capture structural information beyond the fluorophore spatial resolution. To overcome this limitation, here, we developed a hybrid high-speed atomic force microscopy (HS-AFM) and optical tweezers system. To resolve the challenge of incompatible instrumental configurations, we designed a customized optical tweezers system optimized for HS-AFM. Using this platform, we applied external forces to synthetic DNA secondary structures and directly visualized duplex dissociation and spontaneous reannealing upon force release, demonstrating reversible control of folding. We also captured reversible DNA overstretching and transient secondary structure formation in ssDNA. These findings were further analyzed using molecular dynamics simulations and viscoelastic modeling. This integrated approach provides a powerful platform for investigating folding dynamics and force-coupled mechanisms in biomolecules.

DOI： 10.1021/acs.jpclett.5c03517

Enzyme-Directed Assembly of Antiparallel Cellulose II Nanocrystals: Unraveling the Mechanism Beyond Spontaneous Crystallization 査読有り Open Access

Kuga, T; Sunagawa, N; Kobayashi, K; Yamada, H; Imai, T; Uchihashi, T; Igarashi, K

JACS AU 6 巻 ( 1 ) 頁： 154 - 165 2026年1月

記述言語：英語掲載種別：研究論文（学術雑誌）出版者・発行元：Jacs Au

Humans have long utilized cellulose II, known as regenerated cellulose, for fibers like rayon and Cupra and films like cellophane. While cellulose I, found in nature, consists of parallel molecular chains, cellulose II is characterized by the stable arrangement of molecules in an antiparallel orientation. Enzymatic synthesis of cellulose in vitro also affords cellulose II with various morphologies, from monolayer lamellae crystals to gels, but its formation mechanism remains obscure. Here, we demonstrate that cellodextrin phosphorylase (CDP) catalyzes the synthesis and orchestrates the antiparallel self-assembly of cellulose II nanocrystals, exceeding the paradigm of spontaneous crystallization. High-resolution structural analysis reveals CDP’s key role in dictating crystal size and alignment, bridging the gap between enzymatic catalysis and biodirected material architecture. Our research unveils a unique protein-templated assembly process for advanced cellulose materials, paving the way for enzyme-guided construction of next-generation functional nanostructures.

DOI： 10.1021/jacsau.5c00993

Cross-Linker Hydrophobicity Determines Microgel Nanostructures 査読有り

Yamanaka, K; Nishizawa, Y; Iwase, K; Kawane, D; Atarashi, H; Uchihashi, T; Suzuki, D

ACS APPLIED MATERIALS & INTERFACES 17 巻 ( 52 ) 頁： 70793 - 70804 2025年12月

記述言語：英語掲載種別：研究論文（学術雑誌）出版者・発行元：ACS Applied Materials and Interfaces

In aqueous precipitation polymerization, which enables the synthesis of uniformly sized microgels, the monomers are dissolved and the polymer precipitates, leading to nucleation and growth process driven by hydrophobic interactions. Therefore, the (in)compatibility of chemical species used with aqueous solvents is expected to influence the nanostructure of the resulting microgels. In this study, several cross-linkers differing in the number of ethylene glycol units─hence varying in their compatibility with aqueous solvents─were used to synthesize thermoresponsive poly(N-isopropylacrylamide)-based microgels via aqueous precipitation polymerization. The structure and thermoresponsiveness of the resulting microgels were evaluated using dynamic light scattering and temperature-controllable high-speed atomic force microscopy. While microgels cross-linked with hydrophilic cross-linkers exhibited a hierarchically heterogeneous nanostructure, microgels cross-linked with hydrophobic cross-linker showed significant changes in nanostructure with increasing cross-linker raito, changing from a core–shell structure to a spherical structure with rough surface. Kinetic analysis of polymerization revealed that increasing the amount of hydrophobic cross-linker led to a polymerization mechanism other than precipitation polymerization. Similar nanostructures and thermoresponsiveness were also observed with different types of hydrophobic cross-linker, suggesting the universality of these features. In an era where cross-linkers are no longer limited to maintaining structure but are also expected to provide functionality, this study offers guidance for designing microgels with hydrophobic cross-linkers.

DOI： 10.1021/acsami.5c21164

Visualizing the Functional Dynamics of P-Glycoprotein and Its Modulation by Elacridar via High-Speed Atomic Force Microscopy 査読有り Open Access

Kanaoka Y., Hamaguchi-Suzuki N., Nonaka Y., Yamashita S., Miyashita O., Ito A., Ogasawara S., Tama F., Murata T., Uchihashi T.

International Journal of Molecular Sciences 27 巻 ( 1 ) 2025年12月

担当区分：最終著者,　責任著者記述言語：英語掲載種別：研究論文（学術雑誌）出版者・発行元：International Journal of Molecular Sciences

P-glycoprotein (P-gp) is an ATP-driven transporter that effluxes a wide range of xenobiotics from cells, and its overexpression is a primary cause of multidrug resistance (MDR) in cancer. It is well-established that P-gp functions through conformational changes, yet its large-scale structural dynamics at work have been unexplored. Here, we directly visualized single P-gp molecules reconstituted in nanodiscs using high-speed atomic force microscopy (HS-AFM). The HS-AFM movies revealed that P-gp is intrinsically dynamic in its apo state, with its nucleotide-binding domains (NBDs) undergoing large, spontaneous opening and closing motions. However, addition of ATP stabilized a conformation characterized by NBD proximity with a strong tendency toward closure. We then leveraged this dynamic viewpoint to elucidate the relationship between Elacridar’s function and the resulting structural dynamics of P-gp. Elacridar is designed to overcome multidrug resistance (MDR) in cancer and acts as a potent dual inhibitor of both P-gp and the Breast Cancer Resistance Protein (BCRP), effectively blocking the drug efflux function of these transporters. This inhibitor has suggested concentration-dependent function: it is effluxed as a substrate at low concentrations and acts as an inhibitor at high concentrations. Our direct observations revealed that low concentrations induced active dynamics in P-gp, whereas high concentrations severely restricted its motion, leading to a rigid, non-productive state. Our study provides critical insights into how observing molecular motion itself can unravel complex biological mechanisms.

DOI： 10.3390/ijms27010356

Tip-Scan High-Speed Atomic Force Microscopy in Organic Solvent: A Versatile Tool for Visualizing Dynamic Behaviors of Soft-Materials 査読有り

Matsui H., Ganser C., Tamaki K., Liu Q., Chan F.Y., Uchihashi T., Verma P., Sagara Y., Yagai S., Umakoshi T.

Langmuir 42 巻 ( 1 ) 頁： 448 - 454 2025年12月

記述言語：英語掲載種別：研究論文（学術雑誌）出版者・発行元：Langmuir

High-speed atomic force microscopy (HS-AFM) is a powerful tool for video recording the nanoscale dynamics of samples. Its applications have recently expanded to various fields from biology to chemistry, including supramolecular chemistry and polymer science. Furthermore, the development of a stand-alone tip-scan HS-AFM has offered greater flexibility in installation. As it can be mounted on an optical microscope for integration with optical techniques, it holds great promise for supramolecular chemistry and polymer science. However, tip-scan HS-AFM observations in organic solvents have been challenging due to difficulties in holding them with low surface tension. In this study, we achieved tip-scan HS-AFM observations in organic solvents by developing a glass cell. The cell was carefully designed to be compatible with tip-scan HS-AFM operation in organic solvents. Using the cell, we successfully demonstrated tip-scan HS-AFM observations of supramolecular assemblies in various organic systems. In addition, the large and stable design of the cell allowed for a wider variety of organic solvents than the small wells used in conventional HS-AFM. Tip-scan HS-AFM compatible with organic solvents will contribute to investigating the complex dynamics of supramolecular assemblies and polymers in their native organic environments.

DOI： 10.1021/acs.langmuir.5c04454

Light-intensity-dependent out-of-equilibrium processes toward dimensionally distinct nanopolymorphs 査読有り Open Access

Tamaki K., Hanayama H., Datta S., Silly F., Wada Y., Hashizume D., Adachi K., Uchihashi T., Kawano M., Ganser C., Yagai S.

Chem 頁： 102818 - 102818 2025年11月

記述言語：英語掲載種別：研究論文（学術雑誌）出版者・発行元：Chem

Molecular assemblies that form distinct out-of-equilibrium states in response to varying energy inputs represent a promising platform for designing advanced, autonomous adaptive materials capable of flexibly and diversely responding to environmental stimuli. Herein, we describe a supramolecular polymer system that integrates azobenzene photoisomerization with hydrogen-bond-directed supramolecular polymorphism, enabling the formation of distinct out-of-equilibrium states under varied light intensities. t rans isomers of an azobenzene derivative featuring a barbituric acid merocyanine unit self-assemble into lamellar crystals via two-dimensional nanosheet stacking. Ultraviolet light irradiation of a nanosheet dispersion in nonpolar media at different intensities modulates the proportion of cis isomers, eliciting unique out-of-equilibrium states. Specifically, a strong light facilitates the coassembly of trans and cis isomers into one-dimensional nanofibers through hydrogen bond rearrangement, whereas weaker light drives Ostwald ripening, transforming two-dimensional nanosheets into three-dimensional multilayered structures. High-speed atomic force microscopy reveals the intricate dynamic processes driving these transitions.

DOI： 10.1016/j.chempr.2025.102818

Single-molecule dynamics of fibrinogen and bovine serum albumin adsorption on silica nanoparticle coatings observed using high-speed atomic force microscopy 査読有り Open Access

Arachchi, NH; Russo, L; Molino, PJ; Phung, SL; Uchihashi, T; Higgins, MJ

BIOINTERPHASES 20 巻 ( 6 ) 2025年11月

記述言語：英語掲載種別：研究論文（学術雑誌）出版者・発行元：Biointerphases

High-speed atomic force microscopy (HS-AFM) was used to directly visualize the single-molecule adsorption dynamics of fibrinogen (FG) and bovine serum albumin (BSA) on atomically smooth mica and on silica nanoparticle (SiNP) coatings. By capturing the motion of individual proteins against a static background, HS-AFM enables the resolution of key dynamic processes, including surface diffusion, conformational adaptation, binding and unbinding events, and interfacial fluctuations on nanostructured surfaces. The results revealed two distinct, protein-specific adsorption mechanisms on SiNP coatings. BSA adsorbed via strong protein–surface interactions that promoted conformational adaptation and localized shell-like coverage of individual nanoparticles–progressively occupying interparticle interstices but leaving the overall nanoparticle topography visible. In contrast, FG adsorption followed a concentration-dependent, two-stage process; proteins first adsorbing directly to the nanoparticle surface, and at higher coverages, associated via protein–protein interactions, producing a secondary, dynamic, and loosely bound outer layer. This FG protein layer reduced the root-mean-square roughness of the underlying surface from a peak of ∼13.2 to ∼7.8nm while introducing pronounced molecular-level fluctuations at the interface, as inferred from tip-induced smearing in HS-AFM images. These findings demonstrate that the relevant biological interface is not a static substrate, but a dynamic, structurally defined protein layer, whose properties are dictated by both nanoscale surface topography and the characteristics of the adsorbing proteins.

DOI： 10.1116/6.0004849

Lipid-binding ability of Prx2 regulates the structural and functional properties of exosomes from SW480 cells 査読有り

Trang T.N., Yamada R., Umeda K., Yawata S., Sasaki R., Uchihashi T., Ikliptikawati D.K., Kodera N., Ando T., Hazawa M., Konno H.

Biochemical and Biophysical Research Communications 790 巻頁： 152878 - 152878 2025年11月

記述言語：英語掲載種別：研究論文（学術雑誌）出版者・発行元：Biochemical and Biophysical Research Communications

Exosomes are nanoscale extracellular vesicles that play a critical role in intercellular communication, contributing to health maintenance and disease progression. Since Prx2 is often abundant in exosomes, this study investigated how its lipid-binding ability regulates Prx2's localization and functional state on/within exosomes. In SW480 cells, wild-type Prx2 predominantly localized on the exosomal surface via binding to phosphatidylserine (PS). In contrast, a lipid-binding-deficient mutant (MT) was enriched within the lumen, likely due to increased retention in the cytosol during exosome biogenesis. Despite this difference, high-speed atomic force microscopy (HS-AFM) observations revealed no significant difference in exosome size or stiffness between the two, suggesting preserved vesicular architecture. Importantly, PS-binding triggered a functional switch: surface-localized Prx2 lost its peroxidase activity but acquired molecular chaperone activity. Consistently, WT exosomes strongly suppressed protein aggregation in thermal assays, whereas MT exosomes exhibited weaker effects. These findings identify PS binding as a molecular switch that dictates both the localization and function of Prx2, indicating a role for exosome-associated Prx2 in extracellular protein quality control.

DOI： 10.1016/j.bbrc.2025.152878

Enhanced Adsorption and Enzymatic Hydrolysis of Polyethylene Terephthalate by Cutinase with an N-Terminal Hydrophobic Tether 査読有り

Shuvo M.S.R., Ribitsch D., Gübitz G.M., Seno S., Uchihashi T., Onoda A.

ACS Sustainable Chemistry and Engineering 13 巻 ( 42 ) 頁： 17846 - 17855 2025年10月

記述言語：英語掲載種別：研究論文（学術雑誌）出版者・発行元：ACS Sustainable Chemistry and Engineering

The environmental challenges presented by plastic waste, particularly poly(ethylene terephthalate) (PET), necessitate innovative biodegradation strategies. The cutinase from Thermobifida cellulosilytica, Thc_Cut1 (Cut), was site-specifically conjugated with alkyl tethers of varying lengths (C<inf>3</inf>, C<inf>6</inf>, C<inf>9</inf>) through 1H-1,2,3-triazole-4-carbaldehyde (TA4C) derivatives. These conjugations were designed to enhance affinity for PET by adjusting the enzyme’s hydrophobicity. The enzyme kinetic parameters of both conjugated and unconjugated cutinases revealed that the modifications have a minimal impact on catalytic activity. However, a significant improvement in the PET hydrolysis efficiency was observed. Specifically, hexyl and nonyl TA4C-containing cutinase displayed notable increases in terephthalic acid (TPA) release, exceeding the performance of unconjugated cutinase by 65% and 69%, respectively. Scanning electron microscopy and water contact angle measurements confirmed the enhanced erosion and hydrophilicity of the PET surface following the enzyme treatment. Increased enzyme adsorption on the PET surface for C<inf>6</inf>–Cut and C<inf>9</inf>–Cut was validated by X-ray photoelectron spectroscopy. Moreover, high-speed atomic force microscopy demonstrated faster and more stable adsorption of C<inf>6</inf>–Cut and C<inf>9</inf>–Cut on PET surfaces compared with the slower adsorption of unconjugated cutinase. Additionally, molecular dynamics simulations indicate a higher affinity of conjugated cutinase for PET film. These results suggest that conjugating an alkyl tether to the N-terminus strengthens the interaction between cutinase and PET, improving hydrolysis.

DOI： 10.1021/acssuschemeng.5c05212

Controlled Disassembly of Poly(methyl acrylate) Microparticle-Based Polymers using Ethanol-Water Mixtures 査読有り

Kureha, T; Sasaki, Y; Shinozaki, Y; Wakayama, Y; Namba, K; Nishizawa, Y; Minato, H; Watanabe, N; Uchihashi, T; Fujimoto, K; Suzuki, D

LANGMUIR 41 巻 ( 36 ) 頁： 24225 - 24235 2025年9月

記述言語：英語掲載種別：研究論文（学術雑誌）出版者・発行元：Langmuir

Understanding and controlling the degradation conditions of polymers is essential to achieve an environmentally friendly recycling of polymeric materials without deterioration. In this study, we investigated the disassembly of microparticle-based polymer films into the individual microconstituents under mild conditions such as low temperature and aqueous solutions of ethanol. The solvation of the microparticles and the microparticle-assembled structures are key factors for controlling the disassembly of microparticle-based polymers without performing chemical reactions. In particular, according to light-scattering results and molecular-dynamics calculations, the disassembly can be achieved and accelerated under specific conditions including the optimal volume fraction of the aqueous solutions of ethanol as a good solvent for the microparticles. In addition, the disassembly was studied on samples subjected to thermal annealing, which led to changes in the nanostructures of the microparticle-based polymers, as revealed by small-angle X-ray scattering and atomic-force microscopy. The obtained results demonstrate that our microparticle-based concept provides a promising platform for closed-loop materials recycling that can be expected to contribute to alleviating the environmental problems associated with the excessive use of polymeric materials.

DOI： 10.1021/acs.langmuir.5c02004

The Role of Hydrogen Bonding in the Formation and Dynamics of Two Distinct Types of Cyclic Supramolecular Polymers 査読有り

Nodera Yuki, Watanabe Yuichiro, Ishii Yoshiki, Watanabe Go, Ganser Christian, Uchihashi Takayuki, Sugiyasu Kazunori

Angewandte Chemie International Edition 64 巻 ( 38 ) 2025年9月

担当区分：責任著者記述言語：英語

Cyclic polymers are attractive synthetic targets in both polymer chemistry and supramolecular polymer (SP) chemistry; however, there is a gap between these two research fields in terms of the synthetic strategies involved, and achieving a commonality remains elusive. Herein, we describe the formation of two distinct types of cyclic SPs from structurally similar rosette-shaped molecules. The first SP (Type I) is characterized by a uniform diameter and circularity, whereas the other (Type II) has properties reminiscent of flexible cyclic covalent polymers. Remarkably, the only difference in the structures of the two monomers that form these completely distinct cyclic SPs is the absence (Type I) or presence (Type II) of hydrogen-bonding moieties. An all-atom molecular-dynamics simulation suggested that the Type II SP can adopt various “conformations” due to the dynamic nature of its hydrogen bonds, thereby behaving like a flexible covalent polymer. Real-time high-speed atomic-force microscopy (AFM) confirmed the presence of flexibility in the chains of the Type II SP on the observation timescale (several tens of seconds). Our results therefore highlight an example of a similarity between a covalent polymer and a noncovalent SP that should help to bridge the gap between these polymers, leading to new advanced materials.

Integration of Generative Protein Design with Synthetic Porphyrin Assembly: Metal-Responsive Cyclic Assembly of a Bi-Porphyrin Acquisition Designer Protein 査読有り国際誌

Inaba, H; Onoda, H; Uchihashi, T; Oshima, A; Shoji, O

SMALL 21 巻 ( 49 ) 頁： e05625 2025年8月

担当区分：責任著者記述言語：英語掲載種別：研究論文（学術雑誌）出版者・発行元：Small

De novo protein design has been used to create functional protein assemblies, but its reliance on standard amino acids limits the integration of synthetic supramolecular strategies for precise control of molecular assemblies. Herein, an artificial protein assembly that integrates synthetic supramolecular design with de novo protein engineering is presented. Focusing on unit and connection rigidity, the Bi-Porphyrin Acquisition Designer protein (BiPAD), which captures two synthetic porphyrin molecules via state-of-the-art generative protein design to fuse α/β-folded porphyrin-binding motifs, is designed and created. BiPADs captures rigid porphyrins, each bearing an additional metal coordination site, resulting in a metal-responsive cyclic assembly with the intended structure. Furthermore, high-speed atomic force microscopy reveals dynamic structural changes in the BiPAD assembly. This work expands the designability of artificial protein assemblies, paving the way for the synergistic design of functional systems through the integration of protein engineering and synthetic chemistry.

DOI： 10.1002/smll.202505625

Structural basis for the interaction between the bacterial cell division proteins FtsZ and ZapA 査読有り国際誌

Fujita, J; Kasai, K; Hibino, K; Kagoshima, G; Kamimura, N; Tobita, S; Kato, Y; Uehara, R; Namba, K; Uchihashi, T; Matsumura, H

NATURE COMMUNICATIONS 16 巻 ( 1 ) 頁： 5985 - 5985 2025年7月

担当区分：責任著者記述言語：英語掲載種別：研究論文（学術雑誌）出版者・発行元：Nature Communications

Cell division in most bacteria is regulated by the tubulin homolog FtsZ as well as ZapA, a FtsZ-associated protein. However, how FtsZ and ZapA function coordinately has remained elusive. Here we report the cryo-electron microscopy structure of the ZapA-FtsZ complex at 2.73 Å resolution. The complex forms an asymmetric ladder-like structure, in which the double antiparallel FtsZ protofilament on one side and a single protofilament on the other side are tethered by ZapA tetramers. In the complex, the extensive interactions of FtsZ with ZapA cause a structural change of the FtsZ protofilament, and the formation of the double FtsZ protofilament increases electrostatic repulsion. High-speed atomic force microscopy analysis revealed cooperative interactions of ZapA with FtsZ at a molecular level. Our findings not only provide a structural basis for the interaction between FtsZ and ZapA but also shed light on how ZapA binds to FtsZ protofilaments without disturbing FtsZ dynamics to promote cell division.

DOI： 10.1038/s41467-025-60940-w

Microgravity-Assisted Exploration of the Conformational Space of Amyloid β Affected by Tottori-Type Familial Mutation D7N 査読有り国際誌 Open Access

Yagi-Utsumi, M; Yanaka, S; Burton-Smith, RN; Song, CH; Ganser, C; Yamazaki, C; Kasahara, H; Shimazu, T; Uchihashi, T; Murata, K; Kato, K

ACS CHEMICAL NEUROSCIENCE 16 巻 ( 14 ) 頁： 2682 - 2690 2025年6月

記述言語：英語掲載種別：研究論文（学術雑誌）出版者・発行元：ACS Chemical Neuroscience

The amyloid β (Aβ) Tottori variant (D7N) exhibits unique aggregation behaviors and altered fibril formation, posing challenges for structural characterization. To overcome this, the microgravity environment on the International Space Station was employed to study Tottori-type Aβ40 fibril formation and structure. Under Earth gravity, Tottori-type Aβ40 primarily formed nonfibrillar aggregates, hindering detailed structural analysis. In contrast, microgravity significantly enhanced fibril formation and minimized amorphous aggregates. Cryo-electron microscopy revealed two structurally distinct fibril types, each comprising different protomer conformations. In both types, the N-terminal segment was disordered and nor resolved in the density maps. The D7N mutation disrupts the protection of the core by the N-terminal segment often observed in wild-type Aβ40 fibrils, enhancing the hydrophobicity-mediated aggregation propensity. However, microgravity suppressed kinetic traps and facilitated high-quality fibril formation suitable for structural studies that can explore the free energy landscape of Aβ fibril formation. These findings demonstrate the utility of microgravity for studying familial Aβ variants and potentially accelerate our understanding of Aβ aggregation mechanisms in Alzheimer’s disease.

DOI： 10.1021/acschemneuro.5c00217

Construction of a Cyclic Regular-Triangle Trimer of Cytochrome c555 with a Central Hole Using Sortase A 査読有り国際誌 Open Access

Novientri G., Fujiwara K., Mashima T., Matsuura H., Ogata H., Uchihashi T., Fujii S., Sambongi Y., Hirota S.

Chemistry A European Journal 31 巻 ( 27 ) 頁： e202404736 2025年5月

記述言語：英語掲載種別：研究論文（学術雑誌）出版者・発行元：Chemistry A European Journal

Protein-based supramolecules require precise arrangement of building blocks. A regular-triangle trimer (cp-c<inf>555</inf>)<inf>3</inf> has been constructed using an α-helix-inserted-circular permutant (cp-c<inf>555</inf>) of Aquifex aeolicus cytochrome (cyt) c<inf>555</inf>, where the trimers may dissociate to monomers. In this study, we stabilized the regular-triangle structure by constructing a cyclic regular-triangle of three α-helix-linked cyt c<inf>555</inf> molecules using sortase-mediated ligation (SML). Comparing SML using sortase A for six cp-c<inf>555</inf> variant trimers, the variant with GGG at the N-terminus and LPETG at the C-terminus reacted most efficiently. OP-(c<inf>555</inf>)<inf>3</inf> was designed, in which two cyt c<inf>555</inf> molecules were fused using an α-helix, generating a dimer. The cyt c<inf>555</inf> C-terminal region was attached to the N-terminus of the dimer, and the cyt c<inf>555</inf> N-terminal region was attached to the C-terminus of the dimer using the same α-helix. OP-(c<inf>555</inf>)<inf>3</inf> was expressed in Escherichia coli cells, and the termini were connected by SML, forming a cyclic regular-triangle, CL-(c<inf>555</inf>)<inf>3</inf>. CL-(c<inf>555</inf>)<inf>3</inf> showed higher thermostability than (cp-c<inf>555</inf>)<inf>3</inf> and OP-(c<inf>555</inf>)<inf>3</inf>. CL-(c<inf>555</inf>)<inf>3</inf> structural stability was confirmed using high-speed atomic force microscopy. The X-ray crystal structure of CL-(c<inf>555</inf>)<inf>3</inf> showed a cyclic structure and a nanoporous supramolecular assembly. These results demonstrate that a nanoporous supramolecular assembly can be constructed by designing a cyclic molecule with a central hole using SML.

DOI： 10.1002/chem.202404736

Molecular-level insights into the supramolecular gelation mechanism of urea derivative 査読有り国際誌 Open Access

Kimura S., Adachi K., Ishii Y., Komiyama T., Saito T., Nakayama N., Yokoya M., Takaya H., Yagai S., Kawai S., Uchihashi T., Yamanaka M.

Nature Communications 16 巻 ( 1 ) 頁： 3758 - 3758 2025年4月

担当区分：最終著者,　責任著者記述言語：英語掲載種別：研究論文（学術雑誌）出版者・発行元：Nature Communications

Despite being a promising soft material embodied by molecular self-assembly, the formation mechanism of supramolecular gels remains challenging to fully understand. Here we provide molecular to nanoscopic insights into the formation mechanism of gel-forming fibers from a urea derivative. High-speed atomic force microscopy of the urea derivative revealed the presence of a lag phase prior to the formation of supramolecular fibers, suggesting a nucleation process. The fiber growth kinetics differ at both termini of the fiber, indicating a directional hydrogen-bonding motif by the urea units, which is supported by single-crystal X-ray crystallography of a reference compound. Moreover, we observed an intermittent growth pattern of the fibers with repeated elongation and pause phases. This unique behavior can be simulated by a theoretical block-stacking model. A statistical analysis of the concentration-dependent lag time on macroscopic observation of the gelation suggests the presence of a tetrameric or octameric nucleus of the urea molecules.

DOI： 10.1038/s41467-025-59032-6

From bits to biometrics: sustainable hacking of optical storage technologies for atomic force microscopy and medical applications Open Access

Liao, HS; Wang, JH; Hwang, IS; Danzebrink, HU; Uchihashi, T; Huang, KY; Hwu, EET

JAPANESE JOURNAL OF APPLIED PHYSICS 64 巻 ( 4 ) 2025年4月

記述言語：英語掲載種別：研究論文（学術雑誌）出版者・発行元：Japanese Journal of Applied Physics Part 1 Regular Papers and Short Notes and Review Papers

This article showcases how optical pickup units, a type of optical data storage technology, can be sustainably hacked for advanced applications in atomic force microscopy (AFM) and medical diagnostics. The evolution of these technologies from compact discs to Blu-ray is discussed, and their creative applications in high-precision, cost-effective scientific tools are detailed. The transition from data storage to nanoscale imaging has implications for skin nanotexture biometrics, as demonstrated by the example of high-speed dermal AFM for dermatological analysis. Although several technical challenges arise, this approach can have considerable economic and educational benefits and future possibilities, including integration with internet of things and artificial intelligence for stronger functionality. Innovation grounded in hacking can democratize scientific exploration, promote sustainable research, and advance precision medicine, thereby creating a new paradigm for the development of scientific instrumentation.

DOI： 10.35848/1347-4065/adc746

Single-Molecule Imaging of Wood Xylans on Surfaces and Their Interaction with GH11 Xylanase 査読有り国際共著国際誌 Open Access

Schaubeder, JB; Ganser, C; Palasingh, C; Eibinger, M; Nypelö, T; Uchihashi, T; Spirk, S

BIOMACROMOLECULES 26 巻 ( 3 ) 頁： 1639 - 1646 2025年2月

記述言語：英語掲載種別：研究論文（学術雑誌）出版者・発行元：Biomacromolecules

The knowledge of the molecular properties and arrangements of biopolymers in both solid and solution state are essential in the design of sustainable materials and biomedicine as they are decisive for mechanical strength, flexibility, and biodegradability. However, the structure of most biopolymers at charged interfaces can vary considerably, and their time-dependent visualization in liquid-state still remains challenging. In this work, we employed high-speed atomic force microscopy (HS-AFM) to visualize single xylan macromolecules from alkali-extracted birch and beechwood. On negatively charged mica surfaces, they appeared as individual macromolecules but assembled into aggregates on 3-aminopropyltriethoxysilane (APTES) surfaces (AP-mica). Hence, we further investigated the susceptibility to enzymatic degradation using an endoxylanase, which showed that the individual xylan macromolecules remained intact, while larger assemblies on AP-mica degraded over time. We demonstrate that HS-AFM is a powerful tool for understanding the molecular properties and degradation mechanisms of biopolymers. Moreover, by identifying alignment-dependent binding sites, strategies can be developed to ensure the biodegradability of composite materials by intelligent interface design.

DOI： 10.1021/acs.biomac.4c01446

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