Papers - USUI Kenji
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One-Pot Synthesis and Immobilization of Gold Nanoparticles Using Peptidyl Microbeads
Shuhei Yoshida,Koki Yoshida,Taichi Isozaki,Maho Oura,Makoto Ozaki,Takaaki Tsuruoka, Kenji Usui
Molecules 2025 30 ( 8 ) 1689 2025.4
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Development of detection system for lead ions in mixture solutions using UV-Vis measurements with peptide immobilized microbeads
Shuhei Yoshida, Koki Yoshida, Yoshio Hamada, Takaaki Tsuruoka, Kenji Usui
Scientific Reports 15 3249 2025.1
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Complex and Non-sequential Current Signatures of a β-Hairpin Peptide Confined in a Nanopore International coauthorship
Misa Yamaji, Mauro Chinappi, Blasco Morozzo della Rocca, Kenji Usui, Ryuji Kawano
Analytical Chemistry 97 ( 4 ) 2044 - 2051 2025.1
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Development of a CaCO3 Precipitation Method Using a Peptide and Microwaves Generated by a Magnetron Reviewed
Fumihiro Kayamori, Hiroyuki Togashi, Natsumi Endo, Makoto Ozaki, Kan Hirao, Yonejiro Arimoto, Ryuji Osawa, Takaaki Tsuruoka, Takahito Imai, Kin-ya Tomizaki, Tomohiro Umetani, Nobuhiro Nakanishi, Kenji Usui
Processes 12 ( 7 ) 1327 - 1327 2024.6
Publisher：MDPI AG
Microwave applications, such as microwave ovens and mobile phones, are ubiquitous and indispensable in modern society. As the utilization of microwave technology is becoming more widespread, the effects of microwaves on living organisms and physiological processes have received increased attention. This study aimed to investigate the effects of microwaves on calcium carbonate biomineralization as a model biochemical process. A magnetron oscillator was used to generate 2450 MHz microwaves because magnetrons are relatively inexpensive and widespread. We conducted transmission electron microscopy (TEM), atomic force microscopy (AFM), TEM-electron energy-loss spectroscopy (EELS), dynamic light scattering (DLS), and high-performance liquid chromatography (HPLC) measurements to analyze the calcium carbonate precipitates. Our findings showed the formation of string-like precipitates of calcium carbonate upon microwave irradiation from one direction, similar to those obtained using a semiconductor oscillator, as reported previously. This implied that the distribution of the frequency had little effect on the morphology. Furthermore, spherical precipitates were obtained upon microwave irradiation from two directions, indicating that the morphology could be controlled by varying the direction of microwave irradiation. Magnetrons are versatile and also used in large-scale production; thus, this method has potential in medical and industrial applications.
DOI： 10.3390/pr12071327
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Effect of linearly polarized microwaves on nanomorphology of calcium carbonate mineralization using peptides Reviewed
Kenji Usui, Makoto Ozaki, Kan Hirao, Tsubasa Kosaka, Natsumi Endo, Shuhei Yoshida, Shin-ichiro Yokota, Yonejiro Arimoto, Ryuji Osawa, Nobuhiro Nakanishi, Kin-ya Tomizaki, Tomohiro Umetani, Fumihiro Kayamori
Scientific Reports 13 12027 2023.7
Microwaves are used for diverse applications such as mobile phones, ovens, and therapy devices. However, there are few reports on the effects of microwaves on diseases other than cancer, and on physiological processes. Here, we focused on CaCO3 mineralization as a model of biomineralization and attempted to elucidate the effect of microwaves on CaCO3 mineralization using peptides. We conducted AFM, ζ potential, HPLC, ICP-AES, and relative permittivity measurements. Our findings show that microwaves alter the nanomorphology of the CaCO3 precipitate, from sphere-like particles to string-like structures. Furthermore, microwaves have little effect on the mineralization when the mineralization ability of a peptide is high, but a large effect when the precipitation ability is low. Our findings may be applicable to not only the treatment of teeth and bones but also the development of organic–inorganic nanobiomaterials. This methodology can be expanded to other molecular/atomic reactions under various microwave conditions to alter reaction activity parameters.
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Construction of a Method to Design Fibril-Forming Peptides Applied to Biomaterials from 20 Beta-Sheet Peptides by Statistical Analysis Reviewed
Kazuya Iwata, Taisei Terao, Akira Takekawa, Tomohiro Umetani, Kenji Usui
Peptide Science 2022 141 - 142 2023.3
Authorship：Last author, Corresponding author
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De novo design of a nanopore for single-molecule detection that incorporates a β-hairpin peptide. Reviewed International coauthorship International journal
Keisuke Shimizu, Batsaikhan Mijiddorj, Masataka Usami, Ikuro Mizoguchi, Shuhei Yoshida, Shiori Akayama, Yoshio Hamada, Akifumi Ohyama, Kenji Usui, Izuru Kawamura, Ryuji Kawano
Nature nanotechnology 17 ( 1 ) 67 - 75 2022.1
The amino-acid sequence of a protein encodes information on its three-dimensional structure and specific functionality. De novo design has emerged as a method to manipulate the primary structure for the development of artificial proteins and peptides with desired functionality. This paper describes the de novo design of a pore-forming peptide, named SV28, that has a β-hairpin structure and assembles to form a stable nanopore in a bilayer lipid membrane. This large synthetic nanopore is an entirely artificial device for practical applications. The peptide forms multidispersely sized nanopore structures ranging from 1.7 to 6.3 nm in diameter and can detect DNAs. To form a monodispersely sized nanopore, we redesigned the SV28 by introducing a glycine-kink mutation. The resulting redesigned peptide forms a monodisperse pore with a diameter of 1.7 nm leading to detection of a single polypeptide chain. Such de novo design of a β-hairpin peptide has the potential to create artificial nanopores, which can be size adjusted to a target molecule.
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ウォッシャーディスインフェクターのすすぎ排水中に含まれる洗浄剤残留量の測定によるすすぎ性能評価 Reviewed
三軒隼人、藤田敏、川田原瑠勇、武川公、臼井健二、原田陽滋
医療機器学 91 315 - 332 2021.8
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Ozaki, M., Yoshida, S., Tsuruoka, T., Usui, K.
Chemical Communications 57 ( 6 ) 2021
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Ozaki, M., Imai, T., Tsuruoka, T., Sakashita, S., Tomizaki, K.-Y., Usui, K.
Communications Chemistry 4 ( 1 ) 2021
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Miyazaki, H., Hamada, Y., Takaishi, H., Minamino, Y., Ikeda, H., Mekata, H., Takaishi, M., Yamashita, K., Usui, K.
Analyst 145 ( 9 ) 3211 - 3216 2020.5
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Novel purification process for amyloid beta peptide(1-40) Invited Reviewed
Usui, K., Yokota, S.-I., Iwata, K., Hamada, Y.
Processes 8 ( 4 ) 2020
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Miyazaki, H., Takaishi, H., Ikeda, H., Ariumi, H., Hamada, Y., Yamashita, K., Usui, K.
Processes 8 ( 10 ) 2020
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Miyazaki, H., Samejima, Y., Iwata, K., Minamino, Y., Hikida, S., Ariumi, H., Ikeda, H., Hamada, Y., Yamashita, K., Usui, K.
International Journal of Molecular Sciences 21 ( 21 ) 2020
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Effect of tryptophan residues on gold mineralization by a gold reducing peptide Reviewed
Ozaki, M., Yoshida, S., Oura, M., Tsuruoka, T., Usui, K.
RSC Advances 10 ( 66 ) 2020
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Gold-Titania Nanocatalyst Generated by Mineralization Using Two Artificial Peptides with DNA Reviewed
Ozaki Makoto, Tomizaki Kin-Ya, Hamada Yoshio, Usui Kenji
JOURNAL OF PEPTIDE SCIENCE 24 S101 2018.9
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Kenji Usui, Shin-Ichiro Yokota, Makoto Ozaki, Shungo Sakashita, Takahito Imai, Kin-Ya Tomizaki
Protein and Peptide Letters 25 ( 1 ) 42 - 47 2018.4
Joint Work
Authorship：Lead author Publisher：Bentham Science Publishers B.V.
Background: A core sequence (the 9 C-terminal residues) of calcification-associated peptide (CAP-1) isolated from the exoskeleton of the red swamp crayfish was previously shown to control calcium carbonate precipitation with chitin. In addition, a modified core sequence in which the phosphorylated serine at the N terminus is replaced with serine exhibits was also previously shown to alter precipitation characteristics with chitin. Objectives: We focused on calcium carbonate precipitation and attempted to elucidate aspects of the mechanism underlying mineralization. We attempted to evaluate in detail the effects of modifying the N-terminus in the core sequence on calcium carbonate mineralization without chitin. Methods: The peptide modifications included phosphorylation, dephosphorylation, and a free or acetylated Nterminus. The peptides were synthesized manually on Wang resin using the DIPCI-DMAP method for the first residue, and Fmoc solid phase peptide synthesis with HBTU-HOBt for the subsequent residues. Prior to calcium carbonate precipitation, calcium carbonate was suspended in MilliQ water. Carbon dioxide gas was bubbled into the stirred suspension, then the remaining solid CaCO3 was removed by filtration. The concentration of calcium ions in the solution was determined by standard titration with ethylenediaminetetraacetate. Calcium carbonate precipitation was conducted in a micro tube for 3 h at 37°C. We used the micro-scale techniques AFM (atomic force microscopy) and TEM (transmission electron microscopy), and the macro-scale techniques chelate titration, HPLC, gel filtration, CD (circular dichroism) and DLS (dynamic light scattering). Results: We determined the morphologies of the calcium carbonate deposits using AFM and TEM. The pS peptide provided the best control of the shape and size of the calcium carbonate round particles. The acetylated peptides (Ac-S and Ac-pS) provided bigger particles with various shapes. S peptide provided a mixture of bigger particles and amorphous particles. We verified these findings using DLS. All the peptide samples produced nanostructures of the expected size in agreement with the AFM and TEM results. We estimated the abilities of these peptides to precipitate calcium carbonate by determining the residual calcium hydrogen carbonate concentration by standard titration with ethylenediaminetetraacetate after calcium carbonate precipitation. The Ac-pS peptide showed the lowest residual calcium hydrogen carbonate concentration whereas the S peptide showed the highest, suggesting that the precipitating activities of these peptides towards calcium carbonate correlated with peptide net charge. Then the gel filtration results showed a large oligomer peak and a small oligomer/monomer peak for all peptide samples in agreement with the AFM, TEM and DLS results. CD measurements showed that all the peptides formed random-coil-like structures. Thus, we used both macro-and micro-observation techniques such as chelate titration, DLS, AFM and TEM to show that the calcium carbonate precipitating activities of four derivatives of the core sequence of CAP-1 may correlate with the peptide net charge. Conclusion: These peptides mainly act as a catalyst rather than as a binder or component of the calcium carbonate deposits (as a template). On the other hand, the morphologies of the calcium carbonate deposits appeared to be dependent on the ability of the peptide to assemble and act as a template. Consequently, elucidating the relationship between peptide sequence and the ability of the peptide to assemble would be indispensable for controlling precipitate morphologies in the near future. This knowledge would provide important clues for elucidating the relationship between peptide sequence and mineralization ability, including deposit morphology and precipitating activity, for use in nanobiochemistry and materials chemistry research.
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Yuki Tominaga, Kenji Usui, Akiyoshi Hirata, Hiro-O Ito, Kiyoshi Nokihara
Bioorganic and Medicinal Chemistry 26 ( 12 ) 3210 - 3216 2018.4
Joint Work
Authorship：Lead author Publisher：Elsevier Ltd
A fundamental method has been developed focusing on a facile and rapid examination of periodontal disease. Periodontal disease is an oral disease thought to affect 80% of adults, and early detection with treatment is desirable for the improvement of the quality of life. Unfortunately conventional methods are not consistent as the disease is caused by a number of undefined bacteria and detection relies on the skills of the dentist. Thus an objective detection system is required. We have performed an experiment on saliva using a novel biodetection system, designated PepTenChip®. A disease model for saliva was prepared using a specimen from a healthy subject and a mixture of hemoglobin (f-Hb) and lactate dehydrogenase (LDH), which is used as a periodontal disease marker protein with healthy saliva. PepTenChip® is a peptide microarray in which fluorescent labelled structured peptides are immobilized on a novel amorphous carbon substrate. Since the peptides used as capture molecules are fluorescently labelled, labeling of analytes is not necessary. The fluorescence intensity change before and after application of analytes are detected rather than the ON/OFF detection common to conventional microarrays using a set of antigen–antibody. The fluorescence intensity value changes according to the concentration of captured protein allowing the generation of protein fingerprint (PFP) and dendrograms. The present method does not rely on a “one to one” interaction, unlike conventional biodetection, and advantages can be envisaged in the case of an undefined or unknown cause of disease. The statistical analyses, such as multivariate analyses, allow classification of the type of proteins added in saliva as mimetics of disease. PepTenChip® system is useful and convenient for examination of periodontal disease in health care.
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Editorial: Organic-Inorganic Hybrid Materials and Their Applications Reviewed
Kin-ya Tomizaki, Yoshio Hamada, Kenji Usui
Protein & Peptide Letters 25 2 - 3 2018.4
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Peptides for Silica Precipitation: Amino Acid Sequences for Directing Mineralization Reviewed
Makoto Ozaki, Shungo Sakashita, Yoshio Hamada, Kenji Usui
Protein & Peptide Letters 25 ( 1 ) 15 - 24 2018.4
Joint Work
Publisher：Bentham Science Publishers B.V.
Background: Peptides are promising compounds for use in inorganic or organic-inorganic hybrid syntheses (mineralization) and offer several advantages over proteins. Meanwhile, silica-based nanomaterials have been extensively investigated for many years because of their potential application in a diverse range of technologies, including catalysis, sensing, separation, enzyme immobilization, and gene and drug delivery. Considerable progress has been made over the past decade in understanding the molecular mechanisms underpinning biosilicification and the biomimetic synthesis of patterned nanosilica using peptides. Objectives: This mini-review focuses on various peptide sequences, especially short peptide sequences (30 residues or less), for silica mineralization. Methods: We first briefly review early studies on silica mineralization using proteins to provide background information. This is followed by a discussion of promising peptide sequences and attempts to discern the relationship between amino acid sequence, their potential for mineralization, and the properties of the mineral product. Results: The synthetic control of silica mineralization using engineered proteins, such as recombinant silicateins and silaffins, was inspired by silica biomineralization by natural proteins from organisms (sponges, diatoms, and plants). Concurrently, several papers described the utility of well-structured protein assemblies as templates for silica mineralization. These template-directed syntheses of well-structured silica deposits were first conducted using natural proteins or protein assemblies such as collagen fibers and virus hollow protein tubes. Then we reviewed a selection of short peptides (30 residues or less) that had been successfully used for silica mineralization. Almost all peptides developed to date can be sorted by classification like proteins (synthetic control of silica mineralization or utility of templates for silica mineralization): the first class of peptides is used for peptide-directed synthesis, and the second is used for template-directed synthesis after the peptides have assembled and formed nanostructure such as fibers and tubes. The presented peptides were classified and arranged according to the classification. Additionally, we briefly introduced silica mineralization triggered by the combination of short silica-precipitating peptides and template molecules. Conclusion: In this mini-review we focused on various peptide sequences, especially short peptide sequences of 30 residues or less, designed for silica mineralization. The peptides have been used both for peptide-directed silica mineralization and for template-directed silica mineralization. The recent advances in peptide-driven mineralization reviewed here suggest that it will soon be possible to completely control the silica mineralization process using peptides. Mineralization systems using peptides will provide researchers with new tools for controlling various inorganic syntheses and the production of organic-inorganic materials for nanobiochemistry and materials chemistry research.