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Takayuki Miki, Masashi Awa, Yuki Nishikawa, Shigeki Kiyonaka, Masaki Wakabayashi, Yasushi Ishihama, Itaru Hamachi
Zinc signaling and dynamics play significant roles in many physiological responses and diseases. To understand the physiological roles of zinc in detail, comprehensive identification of proteins under high concentration of mobile zinc ion is crucial. We developed a 'conditional proteomics' approach to identify proteins involved in zinc homeostasis based on a chemical proteomic strategy that utilizes designer zinc-responsive labeling reagents to tag such proteins and quantitative mass spectrometry for their identification...
September 12, 2016: Nature Methods
Hajime Shigemitsu, Takahiro Fujisaku, Shoji Onogi, Tatsuyuki Yoshii, Masato Ikeda, Itaru Hamachi
Hydrogelators are small, self-assembling molecules that form supramolecular nanofiber networks that exhibit unique dynamic properties. Development of supramolecular hydrogels that degrade in response to various biomolecules could potentially be used for applications in areas such as drug delivery and diagnostics. Here we provide a synthetic procedure for preparing redox-responsive supramolecular hydrogelators that are used to create hydrogels that degrade in response to oxidizing or reducing conditions. The synthesis takes ∼2-4 d, and it can potentially be carried out in parallel to prepare multiple hydrogelator candidates...
September 2016: Nature Protocols
Maria Aurineide Rodrigues, Leonardo Hamachi, Paulo Tamaso Mioto, Eduardo Purgatto, Helenice Mercier
Guzmania monostachia is an epiphytic heteroblastic bromeliad that exhibits rosette leaves forming water-holding tanks at maturity. Different portions along its leaf blades can display variable degrees of crassulacean acid metabolism (CAM) up-regulation under drought. Since abscisic acid (ABA) can act as an important long-distance signal, we conducted a joint investigation of ontogenetic and drought impacts on CAM intensity and ABA levels in different leaf groups within the G. monostachia rosette. For this, three groups of leaves were analysed according to their position within the mature-tank rosette (i...
August 13, 2016: Plant Physiology and Biochemistry: PPB
Kei Yamaura, Shigeki Kiyonaka, Tomohiro Numata, Ryuji Inoue, Itaru Hamachi
The fast inhibitory actions of γ-aminobutyric acid (GABA) are mainly mediated by GABAA receptors (GABAARs) in the brain. The existence of multiple ligand-binding sites and a lack of structural information have hampered the efficient screening of drugs capable of acting on GABAARs. We have developed semisynthetic fluorescent biosensors for orthosteric and allosteric GABAAR ligands on live cells via coupling of affinity-based chemical labeling reagents to a bimolecular fluorescence quenching and recovery system...
October 2016: Nature Chemical Biology
Shoji Onogi, Hajime Shigemitsu, Tatsuyuki Yoshii, Tatsuya Tanida, Masato Ikeda, Ryou Kubota, Itaru Hamachi
Self-sorted supramolecular nanofibres-a multicomponent system that consists of several types of fibre, each composed of distinct building units-play a crucial role in complex, well-organized systems with sophisticated functions, such as living cells. Designing and controlling self-sorting events in synthetic materials and understanding their structures and dynamics in detail are important elements in developing functional artificial systems. Here, we describe the in situ real-time imaging of self-sorted supramolecular nanofibre hydrogels consisting of a peptide gelator and an amphiphilic phosphate...
August 2016: Nature Chemistry
Koichiro Sueyoshi, Yuka Sumi, Yoshiaki Inoue, Yoko Kuroda, Kumiko Ishii, Hitoshi Nakayama, Kazuhisa Iwabuchi, Yasutaka Kurishita, Hajime Shigemitsu, Itaru Hamachi, Hiroshi Tanaka
BACKGROUND: The activation of polymorphonuclear neutrophils (PMNs) plays an important role in sepsis. Previously, we showed that ATP release and feedback via ATP receptors are essential for PMN activation; however, the dynamics remain poorly understood. Two new fluorescent chemosensors, PMAP-1 and MitoAP-1, were developed to detect ATP in the plasma membrane and mitochondria of living cells, respectively. In this study, we aimed to evaluate ATP localization using these chemosensors in PMNs of sepsis patients...
December 2016: Annals of Intensive Care
Yuki Yasueda, Tomonori Tamura, Alma Fujisawa, Keiko Kuwata, Shinya Tsukiji, Shigeki Kiyonaka, Itaru Hamachi
Protein functions are tightly regulated by their subcellular localization in live cells, and quantitative evaluation of dynamically altered proteomes in each organelle should provide valuable information. Here, we describe a novel method for organelle-focused chemical proteomics using spatially limited reactions. In this work, mitochondria-localizable reactive molecules (MRMs) were designed that penetrate biomembranes and spontaneously concentrate in mitochondria, where protein labeling is facilitated by the condensation effect...
June 22, 2016: Journal of the American Chemical Society
Yuta Hamachi, Moe Katano, Yohei Ogiwara, Norio Sakai
A novel synthesis of α-aminonitriles is described via an indium-catalyzed three-component coupling reaction of alkynes, amines, and trimethylsilyl cyanide (Me3SiCN). Hydroamination of alkynes with a subsequent nucleophilic addition of Me3SiCN resulted in a novel approach to quaternary α-aminonitrile derivatives.
April 1, 2016: Organic Letters
Herbert M Brothers, Eric Bovens, Antonio Bruni, Tanya M Habitz, Tadashi Hamachi, Yuanhua Han, Zhouhua Ji, Joel J Kerbleski, Claude Letouche, Yi Dong Lu, Regis Nguyen, Michelle L Rivard, Xiaoman Qi, Miki Shoji, Ken Tanaka, Ronald E Tecklenburg
A gas chromatography with flame ionization detection (GC-FID) method for analysis of D4, D5, and D6 cyclic siloxanes in silicone emulsions is described. Sample preparation involves breaking the emulsion with methanol and hexanes, and then analyzing the hexanes phase after derivatization with hexamethyldisilazane (HMDS). Silylation is performed to reduce the potential for formation of cyclic siloxanes during the course of the GC analysis. The accuracy of the method was verified by performing analyses on samples spiked with known levels of D4, D5 and D6 and by comparison to a referee method using atmospheric pressure chemical ionization liquid chromatography with mass spectrometry detection (APCI-LC-MS)...
April 8, 2016: Journal of Chromatography. A
Joseph Khoory, Jessica Estanislau, Abdallah Elkhal, Asmae Lazaar, Mark I Melhorn, Abigail Brodsky, Ben Illigens, Itaru Hamachi, Yasutaka Kurishita, Alexander R Ivanov, Sergey Shevkoplyas, Nathan I Shapiro, Ionita C Ghiran
Acute, inflammatory conditions associated with dysregulated complement activation are characterized by significant increases in blood concentration of reactive oxygen species (ROS) and ATP. The mechanisms by which these molecules arise are not fully understood. In this study, using luminometric- and fluorescence-based methods, we show that ligation of glycophorin A (GPA) on human red blood cells (RBCs) results in a 2.1-fold, NADPH-oxidase-dependent increase in intracellular ROS that, in turn, trigger multiple downstream cascades leading to caspase-3 activation, ATP release, and increased band 3 phosphorylation...
2016: PloS One
Masahiro Kitamura, Motoki Akamatsu, Masamitsu Kawanami, Yasushi Furuichi, Takeo Fujii, Mari Mori, Kazushi Kunimatsu, Hidetoshi Shimauchi, Yorimasa Ogata, Matsuo Yamamoto, Taneaki Nakagawa, Shuichi Sato, Koichi Ito, Takefumi Ogasawara, Yuichi Izumi, Kazuhiro Gomi, Kazuhisa Yamazaki, Hiromasa Yoshie, Mitsuo Fukuda, Toshihide Noguchi, Shogo Takashiba, Hidemi Kurihara, Toshihiko Nagata, Takafumi Hamachi, Katsumasa Maeda, Makoto Yokota, Ryuji Sakagami, Yoshitaka Hara, Kazuyuki Noguchi, Toshi Furuuchi, Takashi Sasano, Enyu Imai, Masatoshi Ohmae, Hayuru Koizumi, Mitsuru Watanuki, Shinya Murakami
We investigated the efficacy, safety, and clinical significance of trafermin, a recombinant human fibroblast growth factor (rhFGF)-2, for periodontal regeneration in intrabony defects in Phase III trials. Study A, a multicenter, randomized, double-blind, placebo-controlled study, was conducted at 24 centers. Patients with periodontitis with 4-mm and 3-mm or deeper probing pocket depth and intrabony defects, respectively, were included. A total of 328 patients were randomly assigned (2:1) to receive 0.3% rhFGF-2 or placebo, and 323 patients received the assigned investigational drug during flap surgery...
April 2016: Journal of Bone and Mineral Research: the Official Journal of the American Society for Bone and Mineral Research
Hajime Shigemitsu, Itaru Hamachi
Stimuli-responsive supramolecular assemblies consisting of small molecules are attractive functional materials for biological applications such as drug delivery, medical diagnosis, enzyme immobilization, and tissue engineering. By use of their dynamic and reversible properties, many supramolecular assemblies responsive to a variety of biomolecules have been designed and synthesized. This review focuses on promising strategies for the construction of such dynamic supramolecular assemblies and their functions...
October 2015: Chemistry, An Asian Journal
Takahiro Hayashi, Yuki Yasueda, Tomonori Tamura, Yousuke Takaoka, Itaru Hamachi
A general technique for introducing biophysical probes into selected receptors in their native environment is valuable for the study of their structure, dynamics, function, and molecular interactions. A number of such techniques rely on genetic engineering, which is not applicable for the study of endogenous proteins, and such approaches often suffer from artifacts due to the overexpression and bulky size of the probes/protein tags used. Here we designed novel catalyst-antibody conjugates capable of introducing small chemical probes into receptor proteins such as epidermal growth factor receptor (EGFR) and human epidermal growth factor receptor 2 (HER2) in a selective manner on the surface of living cells...
April 29, 2015: Journal of the American Chemical Society
Ryou Kubota, Itaru Hamachi
Chemical sensing of amino acids, peptides, and proteins provides fruitful information to understand their biological functions, as well as to develop the medical and technological applications. To detect amino acids, peptides, and proteins in vitro and in vivo, vast kinds of chemical sensors including small synthetic binders/sensors, genetically-encoded fluorescent proteins and protein-based semisynthetic biosensors have been intensely investigated. This review deals with concepts, strategies, and applications of protein recognition and sensing using small synthetic binders/sensors, which are now actively studied but still in the early stage of investigation...
July 7, 2015: Chemical Society Reviews
Tatsuyuki Yoshii, Shoji Onogi, Hajime Shigemitsu, Itaru Hamachi
Multicomponent supramolecular hydrogels are constructed for sensitive, naked-eye detection of small-molecule biomarkers. A dendritic self-immolative molecule and the corresponding enzyme as a signal amplification system were stably embedded in a hydrogen peroxide (H2O2)-responsive supramolecular hydrogel (BPmoc-F3), together with other enzymes. The nanostructure and mechanical strength of the hybrid BPmoc-F3 gel were not substantially diminished by incorporation of these multiple components in the absence of target biomarkers, but could be destroyed by addition of the biomarker through the multiple enzymatic and chemical cascade reactions operating in combination within the gel matrix...
March 11, 2015: Journal of the American Chemical Society
Akifumi Hamachi, Tomoya Okuno, Tomoko Imasaka, Yuichiro Kida, Totaro Imasaka
Multiphoton ionization processes were studied for three types of explosives using a line-tunable ultraviolet femtosecond laser. When peroxides such as triacetone triperoxide (TATP) and hexamethylene triperoxide diamine (HMTD) were ionized through a nonresonant two-photon process, a molecular ion was dominantly observed by reducing the excess energy remaining in the ion. However, an aromatic nitro compound such as 2,4,6-trinitrotoluene (TNT) produced large signals arising from molecular and fragment ions by resonant two-photon ionization...
March 3, 2015: Analytical Chemistry
Soonchul Kang, Hui Zheng, Tao Liu, Kohei Hamachi, Shinji Kanegawa, Kunihisa Sugimoto, Yoshihito Shiota, Shinya Hayami, Masaki Mito, Tetsuya Nakamura, Motohiro Nakano, Michael L Baker, Hiroyuki Nojiri, Kazunari Yoshizawa, Chunying Duan, Osamu Sato
Self-assembly of artificial nanoscale units into superstructures is a prevalent topic in science. In biomimicry, scientists attempt to develop artificial self-assembled nanoarchitectures. However, despite extensive efforts, the preparation of nanoarchitectures with superior physical properties remains a challenge. For example, one of the major topics in the field of molecular magnetism is the development of high-spin (HS) molecules. Here, we report a cyanide-bridged magnetic nanocage composed of 18 HS iron(III) ions and 24 low-spin iron(II) ions...
2015: Nature Communications
Shinya Tsukiji, Itaru Hamachi
Introducing nongenetically encoded, synthetic probes into specific proteins is now recognized as a key component in chemical biology. In particular, the ability to chemically modify specific "native" proteins in various contexts from in vitro to cellular systems is of fundamental importance to study biological systems. We developed a protein-labeling technique termed ligand-directed tosyl (LDT) chemistry for this purpose. This method is capable of labeling specific native proteins with diverse synthetic probes with high site specificity and target selectivity without compromising protein function...
2015: Methods in Molecular Biology
Tomonori Tamura, Itaru Hamachi
Catalysts have long played an essential role in organic synthesis and thus hold potential as tools for chemical protein modification. However, there are only a few examples of catalyst-mediated protein labeling under biological conditions because of the difficulty of designing molecular catalysts that work in aqueous environments with high target selectivity and reaction efficiency. To overcome this situation, we have previously developed a new catalyst-based method, termed affinity-guided DMAP (4-dimethylaminopyridine) (AGD) chemistry, for site-specific protein labeling in a target-selective manner using an acyl transfer reaction...
2015: Methods in Molecular Biology
Ippei Takashima, Ryosuke Kawagoe, Itaru Hamachi, Akio Ojida
The concomitant detection of two biological events facilitates the highly selective and sensitive analysis of specific biological functions. In this article, we report an AND logic-gate-type fluorescent probe that can concurrently sense two biological events in living cells: H2 O2 accumulation and acidification. The probe exhibits a unique fluorescence sensing mechanism, in which a xanthene fluorophore is oxidatively transformed to a xanthone derivative by H2 O2 , thereby resulting in a clear dual-emission change...
January 26, 2015: Chemistry: a European Journal
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