Updated on 2024/05/29

写真a

 
YAMASHITA Atsuko
 
Organization
Faculty of Medicine, Dentistry and Pharmaceutical Sciences Professor
Position
Professor
External link

Degree

  • 博士(農学) ( 京都大学 )

Research Interests

  • 構造生物学

  • Structural Biology

Research Areas

  • Life Science / Structural biochemistry

 

Papers

  • Chemical range recognized by the ligand-binding domain in a representative amino acid-sensing taste receptor, T1r2a/T1r3, from medaka fish Reviewed

    Hikaru Ishida, Norihisa Yasui, Atsuko Yamashita

    PLOS ONE   19 ( 3 )   e0300981 - e0300981   2024.3

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    Authorship:Corresponding author   Language:English   Publishing type:Research paper (scientific journal)   Publisher:Public Library of Science (PLoS)  

    Taste receptor type 1 (T1r) proteins are responsible for recognizing nutrient chemicals in foods. In humans, T1r2/T1r3 and T1r1/T1r3 heterodimers serve as the sweet and umami receptors that recognize sugars or amino acids and nucleotides, respectively. T1rs are conserved among vertebrates, and T1r2a/T1r3 from medaka fish is currently the only member for which the structure of the ligand-binding domain (LBD) has been solved. T1r2a/T1r3 is an amino acid receptor that recognizes various l-amino acids in its LBD as observed with other T1rs exhibiting broad substrate specificities. Nevertheless, the range of chemicals that are recognized by T1r2a/T1r3LBD has not been extensively explored. In the present study, the binding of various chemicals to medaka T1r2a/T1r3LBD was analyzed. A binding assay for amino acid derivatives verified the specificity of this protein to l-α-amino acids and the importance of α-amino and carboxy groups for receptor recognition. The results further indicated the significance of the α-hydrogen for recognition as replacing it with a methyl group resulted in a substantially decreased affinity. The binding ability to the protein was not limited to proteinogenic amino acids, but also to non-proteinogenic amino acids, such as metabolic intermediates. Besides l-α-amino acids, no other chemicals showed significant binding to the protein. These results indicate that all of the common structural groups of α-amino acids and their geometry in the l-configuration are recognized by the protein, whereas a wide variety of α-substituents can be accommodated in the ligand binding sites of the LBDs.

    DOI: 10.1371/journal.pone.0300981

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  • Accelerated Molecular Dynamics and AlphaFold Uncover a Missing Conformational State of Transporter Protein OxlT Reviewed

    Jun Ohnuki, Titouan Jaunet-Lahary, Atsuko Yamashita, Kei-ichi Okazaki

    The Journal of Physical Chemistry Letters   15 ( 3 )   725 - 732   2024.1

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    Publishing type:Research paper (scientific journal)   Publisher:American Chemical Society (ACS)  

    DOI: 10.1021/acs.jpclett.3c03052

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  • Overexpression of the flagellar motor protein MotB sensitizes Bacillus subtilis to aminoglycosides in a motility-independent manner. International journal

    Mio Uneme, Kazuya Ishikawa, Kazuyuki Furuta, Atsuko Yamashita, Chikara Kaito

    PloS one   19 ( 4 )   e0300634   2024

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    Language:English   Publishing type:Research paper (scientific journal)  

    The flagellar motor proteins, MotA and MotB, form a complex that rotates the flagella by utilizing the proton motive force (PMF) at the bacterial cell membrane. Although PMF affects the susceptibility to aminoglycosides, the effect of flagellar motor proteins on the susceptibility to aminoglycosides has not been investigated. Here, we found that MotB overexpression increased susceptibility to aminoglycosides, such as kanamycin and gentamicin, in Bacillus subtilis without affecting swimming motility. MotB overexpression did not affect susceptibility to ribosome-targeting antibiotics other than aminoglycosides, cell wall-targeting antibiotics, DNA synthesis-inhibiting antibiotics, or antibiotics inhibiting RNA synthesis. Meanwhile, MotB overexpression increased the susceptibility to aminoglycosides even in the motA-deletion mutant, which lacks swimming motility. Overexpression of the MotB mutant protein carrying an amino acid substitution at the proton-binding site (D24A) resulted in the loss of the enhanced aminoglycoside-sensitive phenotype. These results suggested that MotB overexpression sensitizes B. subtilis to aminoglycosides in a motility-independent manner. Notably, the aminoglycoside-sensitive phenotype induced by MotB requires the proton-binding site but not the MotA/MotB complex formation.

    DOI: 10.1371/journal.pone.0300634

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  • Investigating the Effect of Substituting a Single Cysteine Residue on the Thermal Stability of an Engineered Sweet Protein, Single-Chain Monellin Reviewed

    Kyosuke Ohnuma, Atsuko Yamashita, Norihisa Yasui

    The Protein Journal   2023.9

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    Language:English   Publishing type:Research paper (scientific journal)   Publisher:Springer Science and Business Media LLC  

    Abstract

    Single-chain monellin (SCM) is an engineered protein that links the two chains of monellin, a naturally sweet-tasting protein. This protein is an attractive candidate for use as a sugar replacement in food and beverages and has numerous other applications. Therefore, generating SCM mutants with improved stability is an active area of research to broaden the range of its potential applications. In this study, we focused on the Cys41 residue of SCM, which is a single cysteine residue present at a structurally important position. This residue is often substituted with Ser. However, this substitution may destabilize SCM because Cys41 is buried in the hydrophobic core of the protein. Therefore, we designed mutants that substituted Ala, Val, and Leu for this residue, namely C41A, C41V, and C41L. We characterized these three mutants, SCM C41S, and wild type (WT). Differential scanning fluorimetric analysis revealed that substituting Cys41 with Ala or Val increased the thermal stability of SCM, while substitution with Ser or Leu decreased its stability. Determination of the crystal structures of SCM C41A and C41V mutants revealed that the overall structures and main chain structures around the 41st residue of both mutants were almost identical to the WT. On the other hand, the orientations of the amino acid side chains near the 41st residue differed among the SCM variants. Taken together, our results indicate that substituting Cys41 with Ala or Val increases the stability of SCM and provide insight into the structural basis of this improvement.

    DOI: 10.1007/s10930-023-10154-0

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    Other Link: https://link.springer.com/article/10.1007/s10930-023-10154-0/fulltext.html

  • Direct binding of calmodulin to the cytosolic C-terminal regions of sweet/umami taste receptors Reviewed

    Atsuki Yoshida, Ayumi Ito, Norihisa Yasui, Atsuko Yamashita

    The Journal of Biochemistry   174 ( 5 )   451 - 459   2023.8

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    Authorship:Corresponding author   Language:English   Publishing type:Research paper (scientific journal)   Publisher:Oxford University Press (OUP)  

    Abstract

    Sweet and umami taste receptors recognize chemicals such as sugars and amino acids on their extracellular side and transmit signals into the cytosol of the taste cell. In contrast to ligands that act on the extracellular side of these receptors, little is known regarding the molecules that regulate receptor functions within the cytosol. In this study, we analysed the interaction between sweet and umami taste receptors and calmodulin, a representative Ca2+-dependent cytosolic regulatory protein. High prediction scores for calmodulin binding were observed on the C-terminal cytosolic side of mouse taste receptor type 1 subunit 3 (T1r3), a subunit that is common to both sweet and umami taste receptors. Pull-down assay and surface plasmon resonance analyses showed different affinities of calmodulin to the C-terminal tails of distinct T1r subtypes. Furthermore, we found that T1r3 and T1r2 showed the highest and considerable binding to calmodulin, whereas T1r1 showed weaker binding affinity. Finally, the binding of calmodulin to T1rs was consistently higher in the presence of Ca2+ than in its absence. The results suggested a possibility of the Ca2+-dependent feedback regulation process of sweet and umami taste receptor signaling by calmodulin.

    DOI: 10.1093/jb/mvad060

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    Other Link: https://academic.oup.com/jb/article-pdf/174/5/451/52799861/mvad060.pdf

  • Structure and mechanism of oxalate transporter OxlT in an oxalate-degrading bacterium in the gut microbiota Reviewed

    Titouan Jaunet-Lahary, Tatsuro Shimamura, Masahiro Hayashi, Norimichi Nomura, Kouta Hirasawa, Tetsuya Shimizu, Masao Yamashita, Naotaka Tsutsumi, Yuta Suehiro, Keiichi Kojima, Yuki Sudo, Takashi Tamura, Hiroko Iwanari, Takao Hamakubo, So Iwata, Kei-ichi Okazaki, Teruhisa Hirai, Atsuko Yamashita

    Nature Communications   14 ( 1 )   2023.4

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    Authorship:Corresponding author   Publishing type:Research paper (scientific journal)   Publisher:Springer Science and Business Media LLC  

    Abstract

    An oxalate-degrading bacterium in the gut microbiota absorbs food-derived oxalate to use this as a carbon and energy source, thereby reducing the risk of kidney stone formation in host animals. The bacterial oxalate transporter OxlT selectively uptakes oxalate from the gut to bacterial cells with a strict discrimination from other nutrient carboxylates. Here, we present crystal structures of oxalate-bound and ligand-free OxlT in two distinct conformations, occluded and outward-facing states. The ligand-binding pocket contains basic residues that form salt bridges with oxalate while preventing the conformational switch to the occluded state without an acidic substrate. The occluded pocket can accommodate oxalate but not larger dicarboxylates, such as metabolic intermediates. The permeation pathways from the pocket are completely blocked by extensive interdomain interactions, which can be opened solely by a flip of a single side chain neighbouring the substrate. This study shows the structural basis underlying metabolic interactions enabling favourable symbiosis.

    DOI: 10.1038/s41467-023-36883-5

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    Other Link: https://www.nature.com/articles/s41467-023-36883-5

  • Chloride ions evoke taste sensations by binding to the extracellular ligand-binding domain of sweet/umami taste receptors Reviewed

    Nanako Atsumi, Keiko Yasumatsu, Yuriko Takashina, Chiaki Ito, Norihisa Yasui, Robert F Margolskee, Atsuko Yamashita

    eLife   12   2023.2

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    Authorship:Corresponding author   Publishing type:Research paper (scientific journal)   Publisher:eLife Sciences Publications, Ltd  

    Salt taste sensation is multifaceted: NaCl at low or high concentrations is preferably or aversively perceived through distinct pathways. Cl is thought to participate in taste sensation through an unknown mechanism. Here, we describe Cl ion binding and the response of taste receptor type 1 (T1r), a receptor family composing sweet/umami receptors. The T1r2a/T1r3 heterodimer from the medaka fish, currently the sole T1r amenable to structural analyses, exhibited a specific Cl binding in the vicinity of the amino-acid-binding site in the ligand-binding domain (LBD) of T1r3, which is likely conserved across species, including human T1r3. The Cl binding induced a conformational change in T1r2a/T1r3LBD at sub- to low-mM concentrations, similar to canonical taste substances. Furthermore, oral Cl application to mice increased impulse frequencies of taste nerves connected to T1r-expressing taste cells and promoted their behavioral preferences attenuated by a T1r-specific blocker or T1r3 knock-out. These results suggest that the Cl evokes taste sensations by binding to T1r, thereby serving as another preferred salt taste pathway at a low concentration.

    DOI: 10.7554/eLife.84291

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    Other Link: https://cdn.elifesciences.org/articles/84291/elife-84291-v1.xml

  • Pivotal role for S-nitrosylation of DNA methyltransferase 3B in epigenetic regulation of tumorigenesis. Reviewed International journal

    Kosaku Okuda, Kengo Nakahara, Akihiro Ito, Yuta Iijima, Ryosuke Nomura, Ashutosh Kumar, Kana Fujikawa, Kazuya Adachi, Yuki Shimada, Satoshi Fujio, Reina Yamamoto, Nobumasa Takasugi, Kunishige Onuma, Mitsuhiko Osaki, Futoshi Okada, Taichi Ukegawa, Yasuo Takeuchi, Norihisa Yasui, Atsuko Yamashita, Hiroyuki Marusawa, Yosuke Matsushita, Toyomasa Katagiri, Takahiro Shibata, Koji Uchida, Sheng-Yong Niu, Nhi B Lang, Tomohiro Nakamura, Kam Y J Zhang, Stuart A Lipton, Takashi Uehara

    Nature communications   14 ( 1 )   621 - 621   2023.2

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    DNA methyltransferases (DNMTs) catalyze methylation at the C5 position of cytosine with S-adenosyl-L-methionine. Methylation regulates gene expression, serving a variety of physiological and pathophysiological roles. The chemical mechanisms regulating DNMT enzymatic activity, however, are not fully elucidated. Here, we show that protein S-nitrosylation of a cysteine residue in DNMT3B attenuates DNMT3B enzymatic activity and consequent aberrant upregulation of gene expression. These genes include Cyclin D2 (Ccnd2), which is required for neoplastic cell proliferation in some tumor types. In cell-based and in vivo cancer models, only DNMT3B enzymatic activity, and not DNMT1 or DNMT3A, affects Ccnd2 expression. Using structure-based virtual screening, we discovered chemical compounds that specifically inhibit S-nitrosylation without directly affecting DNMT3B enzymatic activity. The lead compound, designated DBIC, inhibits S-nitrosylation of DNMT3B at low concentrations (IC50 ≤ 100 nM). Treatment with DBIC prevents nitric oxide (NO)-induced conversion of human colonic adenoma to adenocarcinoma in vitro. Additionally, in vivo treatment with DBIC strongly attenuates tumor development in a mouse model of carcinogenesis triggered by inflammation-induced generation of NO. Our results demonstrate that de novo DNA methylation mediated by DNMT3B is regulated by NO, and DBIC protects against tumor formation by preventing aberrant S-nitrosylation of DNMT3B.

    DOI: 10.1038/s41467-023-36232-6

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  • T1R2-mediated sweet sensing in a lizard. Reviewed

    Liang Q, Ko MC, Ng NSR, Borja Reh, Lee JGH, Yamashita A, Nishihara H, Toda Y, Baldwin MW

    Current biology : CB   32 ( 23 )   R1302 - R1303   2022.12

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    Language:English   Publishing type:Research paper (scientific journal)   Publisher:Elsevier BV  

    Sugars are an important class of nutrients found in the flowers and fruits of angiosperms (flowering plants). Although T1R2-T1R3 has been identified as the mammalian sweet receptor, some birds rely on a repurposed T1R1-T1R3 savory receptor to sense sugars. Moreover, as the radiation of flowering plants occurred later than the last common ancestor of amniotes, sugar may not have been an important diet item for amniotes early in evolution, raising the question of whether T1R2-T1R3 is a universal sugar sensor or only a mammalian innovation. Here, using brief-access behavioral tests and functional characterization of taste receptors, we demonstrate that the nectar-taking Madagascar giant day gecko (Phelsuma grandis) can sense sugars through the T1R2-T1R3 receptor. These results reveal the existence of T1R2-based sweet taste in a non-avian reptile, which has important implications for our understanding of the evolutionary history of sugar detection in amniotes.

    DOI: 10.1016/j.cub.2022.10.061

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  • Development of hidden Markov modeling method for molecular orientations and structure estimation from high-speed atomic force microscopy time-series images. Reviewed International journal

    Tomonori Ogane, Daisuke Noshiro, Toshio Ando, Atsuko Yamashita, Yuji Sugita, Yasuhiro Matsunaga

    PLoS computational biology   18 ( 12 )   e1010384   2022.12

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    High-speed atomic force microscopy (HS-AFM) is a powerful technique for capturing the time-resolved behavior of biomolecules. However, structural information in HS-AFM images is limited to the surface geometry of a sample molecule. Inferring latent three-dimensional structures from the surface geometry is thus important for getting more insights into conformational dynamics of a target biomolecule. Existing methods for estimating the structures are based on the rigid-body fitting of candidate structures to each frame of HS-AFM images. Here, we extend the existing frame-by-frame rigid-body fitting analysis to multiple frames to exploit orientational correlations of a sample molecule between adjacent frames in HS-AFM data due to the interaction with the stage. In the method, we treat HS-AFM data as time-series data, and they are analyzed with the hidden Markov modeling. Using simulated HS-AFM images of the taste receptor type 1 as a test case, the proposed method shows a more robust estimation of molecular orientations than the frame-by-frame analysis. The method is applicable in integrative modeling of conformational dynamics using HS-AFM data.

    DOI: 10.1371/journal.pcbi.1010384

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  • An optogenetic assay method for electrogenic transporters using Escherichia coli co‐expressing light‐driven proton pump Reviewed

    Masahiro Hayashi, Keiichi Kojima, Yuki Sudo, Atsuko Yamashita

    Protein Science   30 ( 10 )   2161 - 2169   2021.7

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    Authorship:Last author, Corresponding author   Language:English   Publishing type:Research paper (scientific journal)   Publisher:Wiley  

    DOI: 10.1002/pro.4154

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    Other Link: https://onlinelibrary.wiley.com/doi/full-xml/10.1002/pro.4154

  • A sweet protein monellin as a non-antibody scaffold for synthetic binding proteins Reviewed

    Norihisa Yasui, Kazuaki Nakamura, Atsuko Yamashita

    The Journal of Biochemistry   169 ( 5 )   585 - 599   2021.7

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    Language:English   Publishing type:Research paper (scientific journal)   Publisher:Oxford University Press ({OUP})  

    <title>Abstract</title>
    Synthetic binding proteins that have the ability to bind with molecules can be generated using various protein domains as non-antibody scaffolds. These designer proteins have been used widely in research studies, as their properties overcome the disadvantages of using antibodies. Here, we describe the first application of a phage display to generate synthetic binding proteins using a sweet protein, monellin, as a non-antibody scaffold. Single-chain monellin (scMonellin), in which two polypeptide chains of natural monellin are connected by a short linker, has two loops on one side of the molecule. We constructed phage display libraries of scMonellin, in which the amino acid sequence of the two loops is diversified. To validate the performance of these libraries, we sorted them against the folding mutant of the green fluorescent protein variant (GFPuv) and yeast small ubiquitin-related modifier. We successfully obtained scMonellin variants exhibiting moderate but significant affinities for these target proteins. Crystal structures of one of the GFPuv-binding variants in complex with GFPuv revealed that the two diversified loops were involved in target recognition. scMonellin, therefore, represents a promising non-antibody scaffold in the design and generation of synthetic binding proteins. We termed the scMonellin-derived synthetic binding proteins ‘SWEEPins’.

    DOI: 10.1093/jb/mvaa147

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    Other Link: http://academic.oup.com/jb/article-pdf/169/5/585/38860175/mvaa147.pdf

  • Current pivotal strategies leading a difficult target protein to a sample suitable for crystallographic analysis Invited Reviewed

    Atsuko Yamashita

    Biochemical Society Transactions   48 ( 4 )   1661 - 1673   2020.8

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    Authorship:Lead author, Corresponding author   Publishing type:Research paper (scientific journal)   Publisher:Portland Press Ltd.  

    Crystallographic structural analysis is an essential method for the determination of protein structure. However, crystallization of a protein of interest is the most difficult process in the analysis. The process is often hampered during the sample preparation, including expression and purification. Even after a sample has been purified, not all candidate proteins crystallize. In this mini-review, the current methodologies used to overcome obstacles encountered during protein crystallization are sorted. Specifically, the strategy for an effective crystallization is compared with a pipeline where various expression hosts and constructs, purification and crystallization conditions, and crystallization chaperones as target-specific binder proteins are assessed by a precrystallization screening. These methodologies are also developed continuously to improve the process. The described methods are useful for sample preparation in crystallographic analysis and other structure determination techniques, such as cryo-electron microscopy.

    DOI: 10.1042/BST20200106

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  • Differential scanning fluorimetric analysis of the amino-acid binding to taste receptor using a model receptor protein, the ligand-binding domain of fish T1r2a/T1r3 Reviewed

    Atsuko Yamashita

    Plos One   14   e0218909   2019.10

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    Authorship:Corresponding author   Language:English   Publishing type:Research paper (scientific journal)  

    DOI: 10.1371/JOURNAL.PONE.0218909

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  • Specific modification at the C-terminal lysine residue of the green fluorescent protein variant, GFPuv, expressed in Escherichia coli Reviewed

    Atsuko Yamashita

    Scientific Reports   9   4722   2019.3

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    Authorship:Corresponding author   Language:English   Publishing type:Research paper (scientific journal)  

    DOI: 10.1038/S41598-019-41309-8

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  • うま味受容体細胞外リガンド結合ドメインのX線結晶構造解析 Reviewed

    細谷 麻以子, 山下 敦子

    SPring-8/SACLA利用研究成果集   7   2019.1

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    Authorship:Corresponding author   Language:Japanese  

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  • Structure-function relationships of olfactory and taste receptors Reviewed

    Maik Behrens, Loïc Briand, Claire A. de March, Hiroaki Matsunami, Atsuko Yamashita, Wolfgang Meyerhof, Simone Weyand

    Chemical Senses   43 ( 2 )   81 - 87   2018.2

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    Language:English   Publishing type:Research paper (scientific journal)   Publisher:Oxford University Press  

    The field of chemical senses has made major progress in understanding the cellular mechanisms of olfaction and taste in the past 2 decades. However, the molecular understanding of odor and taste recognition is still lagging far behind and will require solving multiple structures of the relevant full-length receptors in complex with native ligands to achieve this goal. However, the development of multiple complimentary strategies for the structure determination of G proteincoupled receptors (GPCRs) makes this goal realistic. The common conundrum of how multispecific receptors that recognize a large number of different ligands results in a sensory perception in the brain will only be fully understood by a combination of high-resolution receptor structures and functional studies. This review discusses the first steps on this pathway, including biochemical and physiological assays, forward genetics approaches, molecular modeling, and the first steps towards the structural biology of olfactory and taste receptors.

    DOI: 10.1093/chemse/bjx083

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  • A large-scale expression strategy for multimeric extracellular protein complexes using Drosophila S2 cells and its application to the recombinant expression of heterodimeric ligand-binding domains of taste receptor Reviewed

    Atsuko Yamashita, Eriko Nango, Yuji Ashikawa

    PROTEIN SCIENCE   26 ( 11 )   2291 - 2301   2017.11

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    Authorship:Lead author, Corresponding author   Language:English   Publishing type:Research paper (scientific journal)   Publisher:WILEY  

    Many of the extracellular proteins or extracellular domains of plasma membrane proteins exist or function as homo- or heteromeric multimer protein complexes. Successful recombinant production of such proteins is often achieved by co-expression of the components using eukaryotic cells via the secretory pathway. Here we report a strategy addressing large-scale expression of hetero-multimeric extracellular domains of plasma membrane proteins and its application to the extracellular domains of a taste receptor. The target receptor consists of a heterodimer of T1r2 and T1r3 proteins, and their extracellular ligand binding domains (LBDs) are responsible for the perception of major taste substances. However, despite the functional importance, recombinant production of the heterodimeric proteins has so far been unsuccessful. We achieved the successful preparation of the heterodimeric LBD by use of Drosophila S2 cells, which have a high secretory capacity, and by the establishment of a stable high-expression clone producing both subunits at a comparable level. The method overcame the problems encountered in the conventional transient expression of the receptor protein in insect cells using baculovirus or vector lipofection, which failed in the proper heterodimer production because of the biased expression of T1r3LBD over T1r2LBD. The large-scale expression methodology reported here may serve as one of the considerable strategies for the preparation of multimeric extracellular protein complexes.

    DOI: 10.1002/pro.3271

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  • Structural basis for perception of diverse chemical substances by T1r taste receptors Reviewed

    Nipawan Nuemket, Norihisa Yasui, Yuko Kusakabe, Yukiyo Nomura, Nanako Atsumi, Shuji Akiyama, Eriko Nango, Yukinari Kato, Mika K. Kaneko, Junichi Takagi, Maiko Hosotani, Atsuko Yamashita

    NATURE COMMUNICATIONS   8 ( 1 )   15530   2017.5

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    Authorship:Corresponding author   Language:English   Publishing type:Research paper (scientific journal)   Publisher:NATURE PUBLISHING GROUP  

    The taste receptor type 1 (T1r) family perceives 'palatable' tastes. These receptors function as T1r2-T1r3 and T1r1-T1r3 heterodimers to recognize a wide array of sweet and umami (savory) tastes in sugars and amino acids. Nonetheless, it is unclear how diverse tastes are recognized by so few receptors. Here we present crystal structures of the extracellular ligand-binding domains (LBDs), the taste recognition regions of the fish T1r2-T1r3 heterodimer, bound to different amino acids. The ligand-binding pocket in T1r2LBD is rich in aromatic residues, spacious and accommodates hydrated percepts. Biophysical studies show that this binding site is characterized by a broad yet discriminating chemical recognition, contributing for the particular trait of taste perception. In contrast, the analogous pocket in T1r3LBD is occupied by a rather loosely bound amino acid, suggesting that the T1r3 has an auxiliary role. Overall, we provide a structural basis for understanding the chemical perception of taste receptors.

    DOI: 10.1038/ncomms15530

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    Other Link: http://www.nature.com/articles/ncomms1553

  • Taste substance binding elicits conformational change of taste receptor T1r heterodimer extracellular domains Reviewed

    Eriko Nango, Shuji Akiyama, Saori Maki-Yonekura, Yuji Ashikawa, Yuko Kusakabe, Elena Krayukhina, Takahiro Maruno, Susumu Uchiyama, Nipawan Nuemket, Koji Yonekura, Madoka Shimizu, Nanako Atsumi, Norihisa Yasui, Takaaki Hikima, Masaki Yamamoto, Yuji Kobayashi, Atsuko Yamashita

    SCIENTIFIC REPORTS   6   25745   2016.5

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    Authorship:Corresponding author   Language:English   Publishing type:Research paper (scientific journal)   Publisher:NATURE PUBLISHING GROUP  

    Sweet and umami tastes are perceived by T1r taste receptors in oral cavity. T1rs are class C G-protein coupled receptors (GPCRs), and the extracellular ligand binding domains (LBDs) of T1r1/T1r3 and T1r2/T1r3 heterodimers are responsible for binding of chemical substances eliciting umami or sweet taste. However, molecular analyses of T1r have been hampered due to the difficulties in recombinant expression and protein purification, and thus little is known about mechanisms for taste perception. Here we show the first molecular view of reception of a taste substance by a taste receptor, where the binding of the taste substance elicits a different conformational state of T1r2/T1r3 LBD heterodimer. Electron microscopy has showed a characteristic dimeric structure. Forster resonance energy transfer and X-ray solution scattering have revealed the transition of the dimerization manner of the ligand binding domains, from a widely spread to compactly organized state upon taste substance binding, which may correspond to distinct receptor functional states.

    DOI: 10.1038/srep25745

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  • Studies on an Acetylcholine Binding Protein Identify a Basic Residue in Loop G on the beta 1 Strand as a New Structural Determinant of Neonicotinoid Actions Reviewed

    Makoto Ihara, Toshihide Okajima, Atsuko Yamashita, Takuma Oda, Takuya Asano, Mikana Matsui, David B. Sattelle, Kazuhiko Matsuda

    MOLECULAR PHARMACOLOGY   86 ( 6 )   736 - 746   2014.12

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    Authorship:Lead author   Language:English   Publishing type:Research paper (scientific journal)   Publisher:AMER SOC PHARMACOLOGY EXPERIMENTAL THERAPEUTICS  

    Neonicotinoid insecticides target insect nicotinic acetylcholine receptors (nAChRs). Their widespread use and possible risks to pollinators make it extremely urgent to understand the mechanisms underlying their actions on insect nAChRs. We therefore elucidated X-ray crystal structures of the Lymnaea stagnalis acetylcholine binding protein (Ls-AChBP) and its Gln55Arg mutant, more closely resembling insect nAChRs, in complex with a nitromethylene imidacloprid analog (CH-IMI) and desnitro-imidacloprid metabolite (DN-IMI) as well as commercial neonicotinoids, imidacloprid, clothianidin, and thiacloprid. Unlike imidacloprid, clothianidin, and CH-IMI, thiacloprid did not stack with Tyr185 in the wild-type Ls-AChBP, but did in the Gln55Arg mutant, interacting electrostatically with Arg55. In contrast, DN-IMI lacking the NO2 group was directed away from Lys34 and Arg55 to form hydrogen bonds with Tyr89 in loop A and the main chain carbonyl of Trp143 in loop B. Unexpectedly, we found that several neonicotinoids interacted with Lys34 in loop G on the beta 1 strand in the crystal structure of the Gln55Arg mutant. Basic residues introduced into the alpha 7 nAChR at positions equivalent to AChBP Lys34 and Arg55 enhanced agonist actions of neonicotinoids, while reducing the actions of acetylcholine, (-)-nicotine, and DN-IMI. Thus, not only the basic residues in loop D, but also those in loop G determine the actions of neonicotinoids. These novel findings provide new insights into the modes of action of neonicotinoids and emerging derivatives.

    DOI: 10.1124/mol.114.094698

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  • Distinct Human and Mouse Membrane Trafficking Systems for Sweet Taste Receptors T1r2 and T1r3 Reviewed

    Madoka Shimizu, Masao Goto, Takayuki Kawai, Atsuko Yamashita, Yuko Kusakabe

    PLOS ONE   9 ( 7 )   e100425   2014.7

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    The sweet taste receptors T1r2 and T1r3 are included in the T1r taste receptor family that belongs to class C of the G protein-coupled receptors. Heterodimerization of T1r2 and T1r3 is required for the perception of sweet substances, but little is known about the mechanisms underlying this heterodimerization, including membrane trafficking. We developed tagged mouse T1r2 and T1r3, and human T1R2 and T1R3 and evaluated membrane trafficking in human embryonic kidney 293 (HEK293) cells. We found that human T1R3 surface expression was only observed when human T1R3 was coexpressed with human T1R2, whereas mouse T1r3 was expressed without mouse T1r2 expression. A domain-swapped chimera and truncated human T1R3 mutant showed that the Venus flytrap module and cysteine-rich domain (CRD) of human T1R3 contain a region related to the inhibition of human T1R3 membrane trafficking and coordinated regulation of human T1R3 membrane trafficking. We also found that the Venus flytrap module of both human T1R2 and T1R3 are needed for membrane trafficking, suggesting that the coexpression of human T1R2 and T1R3 is required for this event. These results suggest that the Venus flytrap module and CRD receive taste substances and play roles in membrane trafficking of human T1R2 and T1R3. These features are different from those of mouse receptors, indicating that human T1R2 and T1R3 are likely to have a novel membrane trafficking system.

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  • General flexible nature of the cytosolic regions of fungal transient receptor potential (TRP) channels, revealed by expression screening using GFP-fusion techniques Reviewed

    Makoto Ihara, Yoshitaka Takano, Atsuko Yamashita

    PROTEIN SCIENCE   23 ( 7 )   923 - 931   2014.7

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    Transient receptor potential (TRP) channels are members of the voltage gated ion channel superfamily and display the unique characteristic of activation by diverse stimuli. We performed an expression analysis of fungal TRP channels, which possess relatively simple structures yet share the common functional characteristics with the other members, using a green fluorescent protein-based screening methodology. The analysis revealed that all the tested fungal TRP channels were severely digested in their cytosolic regions during expression, implying the common flexibility of this region, as observed in the recent structural analyses of the fungal member, TRPGz. These characteristics are likely to be important for their diverse functions.

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  • Molecular bases of multimodal regulation of a fungal transient receptor potential (TRP) channel Reviewed

    Makoto Ihara, Shin Hamamoto, Yohei Miyanoiri, Mitsuhiro Takeda, Masatsune Kainosho, Isamu Yabe, Nobuyuki Uozumi, Atsuko Yamashita

    Journal of Biological Chemistry   288 ( 21 )   15303 - 15317   2013.5

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    Multimodal activation by various stimuli is a fundamental characteristic of TRP channels. We identified a fungal TRP channel, TRPGz, exhibiting activation by hyperosmolarity, temperature increase, cytosolic Ca2+ elevation, membrane potential, and H2O2 application, and thus it is expected to represent a prototypic multimodal TRP channel. TRPGz possesses a cytosolic C-terminal domain (CTD), primarily composed of intrinsically disordered regions with some regulatory modules, a putative coiled-coil region and a basic residue cluster. The CTD oligomerization mediated by the coiled-coil region is required for the hyperosmotic and temperature increase activations but not for the tetrameric channel formation or other activation modalities. In contrast, the basic cluster is responsible for general channel inhibition, by binding to phosphatidylinositol phosphates. The crystal structure of the presumed coiled-coil region revealed a tetrameric assembly in an offset spiral rather than a canonical coiled-coil. This structure underlies the observed moderate oligomerization affinity enabling the dynamic assembly and disassembly of the CTD during channel functions, which are compatible with the multimodal regulation mediated by each functional module. © 2013 by The American Society for Biochemistry and Molecular Biology, Inc.

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  • GFP-based evaluation system of recombinant expression through the secretory pathway in insect cells and its application to the extracellular domains of class C GPCRs Reviewed

    Yuji Ashikawa, Makoto Ihara, Noriko Matsuura, Yuko Fukunaga, Yuko Kusakabe, Atsuko Yamashita

    PROTEIN SCIENCE   20 ( 10 )   1720 - 1734   2011.10

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    Applications of the GFP-fusion technique have greatly facilitated evaluations of the amounts and qualities of sample proteins used for structural analyses. In this study, we applied the GFP-based sample evaluation to secreted protein expression by insect cells. We verified that a GFP variant, GFPuv, retains proper folding and monodispersity within all expression spaces in Sf9 cells, such as the cytosol, organelles, and even the extracellular space after secretion, and thus can serve as a proper folding reporter for recombinant proteins. We then applied the GFPuv-based system to the extracellular domains of class C G-protein coupled receptors (GPCRs) and examined their localization, folding, and oligomerization upon insect cell expression. The extracellular domain of metabotropic glutamate receptor 1 (mGluR1) exhibited good secreted expression by Sf9 cells, and the secreted proteins formed dimer with a monodisperse hydrodynamic state favorable for crystallization, consistent with the results from previous successful structural analyses. In contrast, the extracellular domains of sweet/umami taste receptors (T1R) almost completely remained in the cell. Notably, the T1R and mGluR1 subfractions that remained in the cellular space showed polydisperse hydrodynamic states with large aggregated fractions, without forming dimers. These results indicated that the proper folding and oligomerization of the extracellular domains of the class C GPCR are achieved through the secretory pathway.

    DOI: 10.1002/pro.707

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  • High-resolution Native-PAGE for membrane proteins capable of fluorescence detection and hydrodynamic state evaluation Reviewed

    Makoto Ihara, Noriko Matsuura, Atsuko Yamashita

    ANALYTICAL BIOCHEMISTRY   412 ( 2 )   217 - 223   2011.5

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    An improved native polyacrylamide gel electrophoresis (PAGE) method capable of evaluating the hydrodynamic states of membrane proteins and allowing in-gel fluorescence detection was established. In this method, bis(alkyl) sulfosuccinate is used to provide negative charges for detergent-solubilized membrane proteins to facilitate proper electrophoretic migration without disturbing their native hydrodynamic states. The method achieved high-resolution electrophoretic separation, in good agreement with the elution profiles obtained by size exclusion chromatography. The applicability of in-gel fluorescence detection for tagged green fluorescent protein (GFP) facilitates the analysis of samples without any purification. This method might serve as a general analytical technique for assessing the folding, oligomerization, and protein complex formation of membrane proteins. (C) 2011 Elsevier Inc. All rights reserved.

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  • A Competitive Inhibitor Traps LeuT in an Open-to-Out Conformation Reviewed

    Satinder K. Singh, Chayne L. Piscitelli, Atsuko Yamashita, Eric Gouaux

    SCIENCE   322 ( 5908 )   1655 - 1661   2008.12

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    Secondary transporters are workhorses of cellular membranes, catalyzing the movement of small molecules and ions across the bilayer and coupling substrate passage to ion gradients. However, the conformational changes that accompany substrate transport, the mechanism by which a substrate moves through the transporter, and principles of competitive inhibition remain unclear. We used crystallographic and functional studies on the leucine transporter ( LeuT), a model for neurotransmitter sodium symporters, to show that various amino acid substrates induce the same occluded conformational state and that a competitive inhibitor, tryptophan ( Trp), traps LeuT in an open- to- out conformation. In the Trp complex, the extracellular gate residues arginine 30 and aspartic acid 404 define a second weak binding site for substrates or inhibitors as they permeate from the extracellular solution to the primary substrate site, which demonstrates how residues that participate in gating also mediate permeation.

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  • Crystal structures of Lymnaea stagnalis AChBP in complex with neonicotinoid insecticides imidacloprid and clothianidin Reviewed

    Makoto Ihara, Toshihide Okajima, Atsuko Yamashita, Takuma Oda, Koichi Hirata, Hisashi Nishiwaki, Takako Morimoto, Miki Akamatsu, Yuji Ashikawa, Shun'Ichi Kuroda, Ryosuke Mega, Seiki Kuramitsu, David B. Sattelle, Kazuhiko Matsuda

    Invertebrate Neuroscience   8 ( 2 )   71 - 81   2008.6

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    Neonicotinoid insecticides, which act on nicotinic acetylcholine receptors (nAChRs) in a variety of ways, have extremely low mammalian toxicity, yet the molecular basis of such actions is poorly understood. To elucidate the molecular basis for nAChR-neonicotinoid interactions, a surrogate protein, acetylcholine binding protein from Lymnaea stagnalis (Ls-AChBP) was crystallized in complex with neonicotinoid insecticides imidacloprid (IMI) or clothianidin (CTD). The crystal structures suggested that the guanidine moiety of IMI and CTD stacks with Tyr185, while the nitro group of IMI but not of CTD makes a hydrogen bond with Gln55. IMI showed higher binding affinity for Ls-AChBP than that of CTD, consistent with weaker CH-π interactions in the Ls-AChBP-CTD complex than in the Ls-AChBP-IMI complex and the lack of the nitro group-Gln55 hydrogen bond in CTD. Yet, the NH at position 1 of CTD makes a hydrogen bond with the backbone carbonyl of Trp143, offering an explanation for the diverse actions of neonicotinoids on nAChRs. © 2008 The Author(s).

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  • Antidepressant binding site in a bacterial homologue of neurotransmitter transporters Reviewed

    Satinder K. Singh, Atsuko Yamashita, Eric Gouaux

    NATURE   448 ( 7156 )   952 - 956   2007.8

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    Sodium-coupled transporters are ubiquitous pumps that harness pre-existing sodium gradients to catalyse the thermodynamically unfavourable uptake of essential nutrients, neurotransmitters and inorganic ions across the lipid bilayer(1). Dysfunction of these integral membrane proteins has been implicated in glucose/galactose malabsorption(2), congenital hypothyroidism(3), Bartter's syndrome(4), epilepsy(5), depression(6), autism(7) and obsessive-compulsive disorder(8). Sodium-coupled transporters are blocked by a number of therapeutically important compounds, including diuretics(9), anticonvulsants(10) and antidepressants(11), many of which have also become indispensable tools in biochemical experiments designed to probe antagonist binding sites and to elucidate transport mechanisms. Steady-state kinetic data have revealed that both competitive(12,13) and noncompetitive(14,15) modes of inhibition exist. Antagonist dissociation experiments on the serotonin transporter (SERT) have also unveiled the existence of a low-affinity allosteric site that slows the dissociation of inhibitors from a separate high-affinity site(16). Despite these strides, atomic-level insights into inhibitor action have remained elusive. Here we screen a panel of molecules for their ability to inhibit LeuT, a prokaryotic homologue of mammalian neurotransmitter sodium symporters, and show that the tricyclic antidepressant (TCA) clomipramine noncompetitively inhibits substrate uptake. Cocrystal structures show that clomipramine, along with two other TCAs, binds in an extracellular-facing vestibule about 11 angstrom above the substrate and two sodium ions, apparently stabilizing the extracellular gate in a closed conformation. Off-rate assays establish that clomipramine reduces the rate at which leucine dissociates from LeuT and reinforce our contention that this TCA inhibits LeuT by slowing substrate release. Our results represent a molecular view into noncompetitive inhibition of a sodium-coupled transporter and define principles for the rational design of new inhibitors.

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  • Structural basis of actin filament capping at the barbed-end: a cryo-electron microscopy study Reviewed

    Atsuko Yamashita

    The EMBO Journal   25 ( 23 )   2006

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    DOI: 10.1038/SJ.EMBOJ.7601395

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  • S3h1-2 Crystal structure of a bacterial homolog of Na^+/Cl^- -dependent neurotransmitter transporters(S3-h1: "Structural Aspects of Channel and Transporter Proteins",Symposia,Abstract,Meeting Program of EABS & BSJ 2006)

    Yamashita Atsuko, Singh Satinder_K, Kawate Toshimitsu, Jin Yan, Gouaux Eric

    Seibutsu Butsuri   46 ( 2 )   S139   2006

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    DOI: 10.2142/biophys.46.S139_4

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  • Crystal structure of a bacterial homologue of Na+/Cl--dependent neurotransmitter transporters Reviewed

    A Yamashita, SK Singh, T Kawate, Y Jin, E Gouaux

    NATURE   437 ( 7056 )   215 - 223   2005.9

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    Na+/Cl--dependent transporters terminate synaptic transmission by using electrochemical gradients to drive the uptake of neurotransmitters, including the biogenic amines, from the synapse to the cytoplasm of neurons and glia. These transporters are the targets of therapeutic and illicit compounds, and their dysfunction has been implicated in multiple diseases of the nervous system. Here we present the crystal structure of a bacterial homologue of these transporters from Aquifex aeolicus, in complex with its substrate, leucine, and two sodium ions. The protein core consists of the first ten of twelve transmembrane segments, with segments 1 - 5 related to 6 - 10 by a pseudo-two-fold axis in the membrane plane. Leucine and the sodium ions are bound within the protein core, halfway across the membrane bilayer, in an occluded site devoid of water. The leucine and ion binding sites are defined by partially unwound transmembrane helices, with main-chain atoms and helix dipoles having key roles in substrate and ion binding. The structure reveals the architecture of this important class of transporter, illuminates the determinants of substrate binding and ion selectivity, and defines the external and internal gates.

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  • 2P222 A new image analysis to determine three dimensional structures of ends of actin filaments from cryo electron micrographs

    Narita A., Yamashita A., Maeda Y.

    Seibutsu Butsuri   45   S175   2005

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    DOI: 10.2142/biophys.45.S175_2

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  • Capping protein binding to actin in yeast: biochemical mechanism and physiological relevance Reviewed International journal

    Kim, K, Yamashita, A, Wear, MA, Maeda, Y, Cooper, JA

    Journal of Cell Biology   164 ( 4 )   567 - 80   2004

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    The mechanism by which capping protein (CP) binds barbed ends of actin filaments is not understood, and the physiological significance of CP binding to actin is not defined. The CP crystal structure suggests that the COOH-terminal regions of the CP alpha and beta subunits bind to the barbed end. Using purified recombinant mutant yeast CP, we tested this model. CP lacking both COOH-terminal regions did not bind actin. The alpha COOH-terminal region was more important than that of beta. The significance of CP's actin-binding activity in vivo was tested by determining how well CP actin-binding mutants rescued null mutant phenotypes. Rescue correlated well with capping activity, as did localization of CP to actin patches, indicating that capping is a physiological function for CP. Actin filaments of patches appear to be nucleated first, then capped with CP. The binding constants of yeast CP for actin suggest that actin capping in yeast is more dynamic than in vertebrates.

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  • Enzyme Structure Prior to Reaction Initiation Capturing with Laue Diffraction and its Reaction Mechanism : Tropinone Reductase-II-Substrate Complexes

    YAMASHITA Atsuko, KATO Hiroaki

    X-RAYS   45 ( 6 )   371 - 377   2003.12

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    To understand the catalytic mechanism of an enzyme, it is crucial to determine the crystallographic structures corresponding to the individual reaction steps. Here we report two crystal structures of enzyme-substrates complexes prior to reaction initiation : tropinone reductase-II (TR-II) -NADPH and TR-II-NADPH-tropinone complexes, determined from the identical crystals. A combination of two kinetic crystallographic techniques, a continuous flow of the substrates and Laue diffraction measurement, enabled us to capture the transit structures prior to the reaction proceeding. A structure comparison of enzyme-substrates complex elucidated in this study with the enzyme-products complex in our previous study indicates that one of the substrates, tropinone, is rotated relative to the product so as to make the spatial organization in the active site favorable for the reaction to proceed. Side chains of the residues in the active site also alter their conformations to keep the complementarily of the space for the substrate or the product and to assist the rotational movement.

    DOI: 10.5940/jcrsj.45.371

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  • Crystal structure of CapZ: structural basis for actin filament barbed end capping International journal

    A. Yamashita

    The EMBO Journal   22 ( 7 )   1529 - 1538   2003.4

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    Capping protein, a heterodimeric protein composed of alpha and beta subunits, is a key cellular component regulating actin filament assembly and organization. It binds to the barbed ends of the filaments and works as a 'cap' by preventing the addition and loss of actin monomers at the end. Here we describe the crystal structure of the chicken sarcomeric capping protein CapZ at 2.1 A resolution. The structure shows a striking resemblance between the alpha and beta subunits, so that the entire molecule has a pseudo 2-fold rotational symmetry. CapZ has a pair of mobile extensions for actin binding, one of which also provides concomitant binding to another protein for the actin filament targeting. The mobile extensions probably form flexible links to the end of the actin filament with a pseudo 2(1) helical symmetry, enabling the docking of the two in a symmetry mismatch.

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  • How capping protein binds the barbed end of the actin filament Reviewed International journal

    Wear, MA, Yamashita, A, Kim, K, Maeda, Y, Cooper, JA

    Current Biology   13 ( 17 )   1531 - 7   2003

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    Cytoskeletal filaments are often capped at one end, regulating assembly and cellular location. The actin filament is a right-handed, two-strand long-pitch helix. The ends of the two protofilaments are staggered in relation to each other, suggesting that capping could result from one protein binding simultaneously to the ends of both protofilaments. Capping protein (CP), a ubiquitous alpha/beta heterodimer in eukaryotes, tightly caps (K(d) approximately 0.1-1 nM) the barbed end of the actin filament (the end favored for polymerization), preventing actin subunit addition and loss. CP is critical for actin assembly and actin-based motility in vivo and is an essential component of the dendritic nucleation model for actin polymerization at the leading edge of cells. However, the mechanism by which CP caps actin filaments is not well understood. The X-ray crystal structure of CP has inspired a model where the C termini ( approximately 30 amino acids) of the alpha and beta subunits of CP are mobile extensions ("tentacles"), and these regions are responsible for high-affinity binding to, and functional capping of, the barbed end. We tested the tentacle model in vitro with recombinant mutant CPs. Loss of both tentacles causes a complete loss of capping activity. The alpha tentacle contributes more to capping affinity and kinetics; its removal reduces capping affinity by 5000-fold and the on-rate of capping by 20-fold. In contrast, removal of the beta tentacle reduced the affinity by only 300-fold and did not affect the on-rate. These two regions are not close to each other in the three-dimensional structure, suggesting CP uses two independent actin binding tentacles to cap the barbed end. CP with either tentacle alone can cap, as can the isolated beta tentacle alone, suggesting that the individual tentacles interact with more than one actin subunit at a subunit interface at the barbed end.

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  • Structure of the core domain of human cardiac troponin in the Ca2+-saturated form Reviewed

    Atsuko Yamashita

    Nature   424 ( 6944 )   2003

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    DOI: 10.1038/NATURE01780

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  • Capturing Enzyme Structure Prior to Reaction Initiation: Tropinone Reductase-II−Substrate Complexes Reviewed

    Atsuko Yamashita

    Biochemistry   42 ( 19 )   2003

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    DOI: 10.1021/BI0272712

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  • Crystal structure of the C-terminal half of tropomodulin and structural basis of actin filament pointed-end capping Reviewed International journal

    Krieger, I, Kostyukova, A, Yamashita, A, Nitanai, Y, Maeda, Y

    Biophysical Journal   83 ( 5 )   2716 - 25   2002

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    Tropomodulin is the unique pointed-end capping protein of the actin-tropomyosin filament. By blocking elongation and depolymerization, tropomodulin regulates the architecture and the dynamics of the filament. Here we report the crystal structure at 1.45-A resolution of the C-terminal half of tropomodulin (C20), the actin-binding moiety of tropomodulin. C20 is a leucine-rich repeat domain, and this is the first actin-associated protein with a leucine-rich repeat. Binding assays suggested that C20 also interacts with the N-terminal fragment, M1-M2-M3, of nebulin. Based on the crystal structure, we propose a model for C20 docking to the actin subunit at the pointed end. Although speculative, the model is consistent with the idea that a tropomodulin molecule competes with an actin subunit for a pointed end. The model also suggests that interactions with tropomyosin, actin, and nebulin are all possible sources of influences on the dynamic properties of pointed-end capping by tropomodulin.

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  • 1E1500 Crystal structures of Troponin ternary complexes and insights into regulation of muscle contraction

    Takeda S., Yamashita A., Maeda K., Maeda Y.

    Seibutsu Butsuri   42 ( 2 )   S32   2002

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    DOI: 10.2142/biophys.42.S32_4

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  • 2B1415 Crystal structure and actin-binding mechanism of actin filament capping protein CapZ

    Yamashita A., Maeda K., Maeda Y.

    Seibutsu Butsuri   42 ( 2 )   S100   2002

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    DOI: 10.2142/biophys.42.S100_4

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  • Strategy for Crystallization and structural analysis of muscle regulatory protein complex

    Takeda S., Yamashita A., Maeda Y.

    Seibutsu Butsuri   41   S36   2001

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  • Crystallographic structure analysis of actin capping protein Capz

    Yamashita A., Maeda K., Maeda Y.

    Seibutsu Butsuri   41   S60   2001

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    DOI: 10.2142/biophys.41.S60_4

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  • Crystallization and preliminary x-ray analysis of troponin ternary complex

    Takeda S., Yamashita A., Maeda K., Kreiger Inna, Maeda Y.

    Seibutsu Butsuri   40   S139   2000

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    DOI: 10.2142/biophys.40.S139_1

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  • Structure of Tropinone Reductase-II Complexed with NADP + and Pseudotropine at 1.9 Å Resolution: Implication for Stereospecific Substrate Binding and Catalysis Reviewed

    Atsuko Yamashita

    Biochemistry   38 ( 24 )   1999

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    DOI: 10.1021/BI9825044

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  • Crystal structures of two tropinone reductases: Different reaction stereospecificities in the same protein fold Reviewed

    Nakajima, K, Yamashita, A, Akama, H, Nakatsu, T, Kato, H, Hashimoto, T, Oda, J, Yamada, Y

    Proceedings of the National Academy of Sciences of the United States of America   95 ( 9 )   4876 - 4881   1998

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    DOI: 10.1073/pnas.95.9.4876

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  • Crystallization and preliminary crystallographic study of tropinone reductase II from Datura stramonium Reviewed

    Atsuko Yamashita

    Acta Crystallographica Section D Structural Biology   54   1998

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    DOI: 10.1107/S0907444998005782

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  • Direct interaction between mitochondrial succinate-ubiquinone and ubiquinol-cytochrome c oxidoreductases probed by sensitivity to quinone-related inhibitors Reviewed

    Yamashita, A, Miyoshi, H, Hatano, T, Iwamura, H

    Journal of Biochemistry   120 ( 2 )   377 - 384   1996

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    The electron-transfer activities of bovine heart rnitochondrial complexes I, II, and III, but not complex IV, were simultaneously inhibited by 2-alkyl-4, 6-dinitrophenols to a different extent. The extent of inhibition of NADH and succinate oxidase activities by dinitrophenols was compared with that of individual complex activities using submitochondrial particles. The extent of inhibition of succinate oxidase activity by 1-methyipropyl and 1-methylbutyl derivatives was much larger than that of NADH oxidase activity. This large inhibition of succinate oxidase activity seemed not to be explainable by the extent of inhibition of individual complex activities (i.e., complexes II and III activities), based upon the homogeneous ubiquinone pool model. On the other hand, other dinitrophenols (n-propyl, 1-methylpentyl, 1-methylhexyl, and tert-butyl derivatives) very similar to the above compounds did not elicit such anomalous inhibitory action, indicating that the action of 1-methylpropyl and 1-methylbutyl derivatives is highly specific to their structure. The anomalous inhibition by these two compounds was also observed with the isolated succinate-cytochrome c oxidoreductase, in which there is no ubiquinone pool behavior [Rich, P. R. (1984) Biochim. Biophys. Acta 768, 53-79]. However, when the succinate-cytochrome c reductase of which the activity had been partially restored by adding phospholipid and exogenous quinone to the phospholipid- and ubiquinone-depleted succinate-cytochrome c reductase was assayed, the anomalous inhibitory action of interest was undetectable. These results indicated that electron-transfer between complexes II and III, which is mediated not only by free-form, but also by protein-bound ubiquinone, occurs in the mitochondrial membrane. The fact that the anomalous inhibition of succinate oxidase activity of submitochondrial particles was sensitive to changes in the external osmotic pressure which affected the total area of the particle supports this notion.

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  • おいしさの科学とフードテック最前線

    都甲, 潔( Role: Contributor ,  第2章「味覚受容体の構造と働き」)

    シーエムシー出版  2022.8  ( ISBN:9784781316758

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    Total pages:xii, 435p   Language:Japanese

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  • 膜タンパク質工学ハンドブック

    津本, 浩平, 浜窪, 隆雄( Role: Contributor ,  第1編第2章第9節「ポリアクリルアミドゲル電気泳動法を利用した膜タンパク質の性状解析」, 第3編第10章「味覚受容体タンパク質を利用した味物質評価法開発の可能性」)

    エヌ・ティー・エス  2020.4  ( ISBN:9784860435370

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    Total pages:i, 5, 14, 538, 18p, 図版42p   Language:Japanese

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  • 口・鼻・耳の感覚メカニズムと応用技術

    ( Role: Contributor ,  第1章・第2節「味覚」)

    S&T出版  2018  ( ISBN:9784907002701

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  • 膜タンパク質構造研究

    ( Role: Contributor ,  「神経伝達物質トランスポーターホモログ」)

    化学同人  2013  ( ISBN:9784759815610

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  • 化学受容の科学: 匂い・味・フェロモン 分子から行動まで

    ( Role: Contributor ,  12章「化学感覚受容体の構造生物学」)

    化学同人  2012  ( ISBN:4759815015

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  • 腸内細菌叢と尿路結石の関係は?−尿路結石形成に影響する腸内シュウ酸分解菌と同菌が有するシュウ酸輸送体の構造と機能 Invited

    山下敦子

    臨床泌尿器科   78 ( 3 )   224 - 230   2024.3

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  • 腸内シュウ酸分解菌で働くシュウ酸輸送体の分子メカニズム Invited

    山下敦子, 岡崎圭一

    生物物理   64 ( 1 )   25 - 27   2024.2

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  • 放射光で視た甘味・うま味受容~味覚受容体T1rの構造解析~ Invited Reviewed

    山下敦子

    SPring-8/SACLA 利用者情報   28 ( 4 )   344 - 348   2023.11

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    Authorship:Lead author   Language:Japanese   Publishing type:Article, review, commentary, editorial, etc. (bulletin of university, research institution)  

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  • 食塩は甘味やうま味の受容体でも感知されている Invited

    山下敦子

    明日の食品産業   ( 539 )   28 - 31   2023.9

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  • 甘味受容体・うま味受容体を介した塩化物イオンの感知 Invited

    山下敦子

    バイオサイエンスとインダストリー   81 ( 5 )   406 - 407   2023.9

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  • 味覚受容体の構造と働き Invited

    山下敦子

    milsil   14 ( 6 )   6 - 8   2021.11

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  • 受容体タンパク質を用いた味覚感知初反応の解析と応用 Invited

    山下敦子, 芦川雄二, 南後恵理子, 安井典久

    バイオサイエンスとインダストリー   79 ( 2 )   86 - 89   2021.3

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    J-GLOBAL

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  • 微生物TRPチャネルの機能と役割

    魚住信之, 山下敦子

    医学のあゆみ   270   970 - 976   2019.9

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  • 食行動と栄養摂取をむすぶ味覚受容体による味分子認識 Invited

    山下 敦子

    実験医学   37   531 - 535   2019.3

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  • 味覚受容体細胞外領域ヘテロ二量体の発現・精製および性状解析(Expression, purification, and characterization of the entire heterodimeric extracellular regions of fish taste receptor)

    Maruhashi Hiroki, Noshiro Daisuke, Yasui Norihisa, Ando Toshio, Yamashita Atsuko

    生物物理   57 ( Suppl.1-2 )   S313 - S313   2017.8

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  • X 線小角散乱法による味覚受容体リガンド結合ドメインの味物質結合に伴う構造変化の解析 Invited

    山下 敦子

    分子研レターズ   75   28 - 29   2017.3

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  • GPCRの機能構造とその形成 Invited

    山下 敦子

    医学のあゆみ   256   365 - 371   2016.1

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  • Taste substance binding to the ligand-binding domains of T1r taste receptor heterodimer

    Yamashita Atsuko, Nango Eriko, Akiyama Shuji, Maki-Yonekura Saori, Ashikawa Yuji, Kusakabe Yuko, Krayukhina Elena, Maruno Takahiro, Uchiyama Susumu, Nuemket Nipawan, Yonekura Koji, Shimizu Madoka, Atsumi Nanako, Yasui Norihisa, Hikima Takaaki, Yamamoto Masaki, Kobayashi Yuji

    Chemical Senses   41 ( 9 )   E176 - E177   2016

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    Web of Science

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  • Role of the C-terminal region of T1r3 in the membrane trafficking of taste receptor T1r2/T1r3

    Atsuko Yamashita

    Chemical Senses   2016

  • TRPチャネルの構造から考える多様な刺激応答 Invited

    山下 敦子

    生化学   86 ( 4 )   513 - 517   2014.4

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  • TRPチャネルがさまざまな刺激に応答できる仕組み

    伊原誠, 山下敦子

    化学と生物   52 ( 1 )   48 - 53   2014

  • From stills to movies-attempts with protein X-ray crystallography

    YAMASHITA Atsuko

    Folia Pharmacologica Japonica   141 ( 5 )   235 - 239   2013

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    Language:Japanese   Publisher:The Japanese Pharmacological Society  

    DOI: 10.1254/fpj.141.235

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    Other Link: https://jlc.jst.go.jp/DN/JALC/10018417660?from=CiNii

  • 膜タンパク質構造解析の新展開

    山下 敦子

    ファルマシア   49 ( 8 )   753 - 757   2013

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    Language:Japanese   Publisher:公益社団法人 日本薬学会  

    DOI: 10.14894/faruawpsj.49.8_753

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  • Membrane trafficking of T1r3 taste receptor

    KUSAKABE Yuko, SHINDO Yumiko, KAWAI Takayuki, YAMASHITA Atsuko

    The Japanese journal of taste and smell research   19 ( 3 )   297 - 298   2012.12

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  • 基質と阻害剤のあいだ:神経伝達物質トランスポーターホモログLeuTの結晶構造から

    山下敦子

    細胞工学   31   527 - 528   2012

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  • Structure basis of a regulatory module of TRP channel homolog

    Atsuko Yamashita, Makoto Ihara

    JOURNAL OF PHARMACOLOGICAL SCIENCES   118   12P - 12P   2012

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  • FN-PAGE法:膜タンパク質の性状の簡便かつ迅速な解析法

    伊原誠, 山下敦子

    実験医学   29   2291 - 2297   2011

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  • 神経伝達物質トランスポーターホモログの結晶構造から考える輸送機構と阻害剤作用機構

    山下敦子

    遺伝子医学MOOK19号「トランスポートソーム:生体膜輸送機構の全体像に迫る−基礎,臨床・創薬応用研究の最新成果」   44 - 50   2011

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  • Membrane-Protein Crystallography and Potentiality for Drug Design : An Example from Neurotransmitter Transporter Homolog LeuT

    YAMASHITA Atsuko

    X-RAYS   52 ( 1 )   76 - 80   2010

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    Language:Japanese   Publisher:The Crystallographic Society of Japan  

    Structure-based drug design for membrane proteins is far behind that for soluble proteins due to difficulty in crystallographic structure determination, despite the fact that about 60% of FDA-approved drugs target membrane proteins located at the cell surface. Stable homologs for a membrane protein of interest, such as prokaryotic neurotransmitter transporter homolog LeuT, might enable cooperative analyses by crystallography and functional assays, provide useful information for functional mechanisms, and thus serve as important probes for drug design based on mechanisms as well as structures.

    DOI: 10.5940/jcrsj.52.76

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  • Inhibition mechanism on neurotransmitter transporter homolog revealed by crystallographic analyses

    Atsuko Yamashita

    Journal of Pharmacological Sciences   2010

  • New Membrane Protein Structures

    YAMASHITA Atsuko

    51 ( 1 )   100 - 101   2009.2

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  • 真核生物由来膜蛋白質の結晶化支援法の開発—GFP融合技術を用いた結晶化能判定系と酵母による蛋白質生産系

    加藤博章, 山下敦子, 江川響子, 崎山慶太

    蛋白質核酸酵素   54   1461 - 1467   2009

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  • 培養細胞を用いたタンパク質発現のリアルタイム測定法

    芦川雄二, 伊原誠, 山下敦子

    日本蛋白質科学会年会プログラム・要旨集   8th   98   2008.5

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    J-GLOBAL

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  • Time-resolved X-ray crystallographic analysis on tropinone reductase-II using Laue diffraction method

    Atsuko Yamashita

    Seikagaku   2005

  • Structural basis for actin filament capping and targeting by CapZ

    A Yamashita, K Maeda, Y Maeda

    BIOPHYSICAL JOURNAL   84 ( 2 )   257A - 257A   2003.2

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  • Crystal structure of troponin ternary complex

    S Takeda, A Yamashita, K Maeda, Y Maeda

    BIOPHYSICAL JOURNAL   82 ( 1 )   170A - 170A   2002.1

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  • Crystal structure of actin capping protein CapZ

    A Yamashita, K Maeda, Y Maeda

    BIOPHYSICAL JOURNAL   82 ( 1 )   384A - 384A   2002.1

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    Web of Science

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  • Crystal structure of the C-terminal half of tropomodulin: A leucine-rich repeat domain caps actin filament

    Krieger, I, A Kostyukova, A Yamashita, Y Nitanai, Y Maeda

    BIOPHYSICAL JOURNAL   82 ( 1 )   384A - 384A   2002.1

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Awards

  • 長瀬研究振興賞

    2023   公益財団法人 長瀬科学技術振興財団  

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  • 生物学研究奨励賞

    2016   公益財団法人 両備檉園記念財団  

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  • 文部科学大臣表彰 若手科学者賞

    2009  

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    Country:Japan

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