Updated on 2021/12/16

写真a

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

Research Interests

  • 構造生物学

  • Structural Biology

Research Areas

  • Life Science / Structural biochemistry

 

Papers

  • 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   2021.10

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    Authorship:Last author, Corresponding author   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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    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

    Yoshida, Takashi, Yasui, Norihisa, Kusakabe, Yuko, Ito, Chiaki, Akamatsu, Miki, Yamashita, Atsuko

    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

    Nakatani, Takahiro, Yasui, Norihisa, Tamura, Issei, Yamashita, Atsuko

    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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    Language:English   Publishing type:Research paper (scientific journal)   Publisher:PUBLIC LIBRARY SCIENCE  

    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.

    DOI: 10.1371/journal.pone.0100425

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

    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.

    DOI: 10.1002/pro.2474

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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.

    DOI: 10.1074/jbc.M112.434795

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

    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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    Authorship:Corresponding author   Language:English   Publishing type:Research paper (scientific journal)   Publisher:ACADEMIC PRESS INC ELSEVIER SCIENCE  

    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.

    DOI: 10.1016/j.ab.2011.01.038

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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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    Language:English   Publishing type:Research paper (scientific journal)   Publisher:AMER ASSOC ADVANCEMENT SCIENCE  

    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.

    DOI: 10.1126/science.1166777

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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).

    DOI: 10.1007/s10158-008-0069-3

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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.

    DOI: 10.1038/nature06038

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

    Narita, Akihiro, Takeda, Shuichi, Yamashita, Atsuko, Maeda, Yuichiro

    Embo Journal   25 ( 23 )   2006

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    DOI: 10.1038/sj.emboj.7601395

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

    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.

    DOI: 10.1038/nature03978

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

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

    Journal of Cell Biology   164 ( 4 )   2004

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    DOI: 10.1083/jcb.200308061

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

    A. Yamashita

    The EMBO Journal   22 ( 7 )   1529 - 1538   2003.4

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    Authorship:Lead author, Corresponding author   Publishing type:Research paper (scientific journal)   Publisher:Wiley  

    DOI: 10.1093/emboj/cdg167

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  • Capturing enzyme structure prior to reaction initiation: Tropinone reductase-II-substrate complexes Reviewed

    Yamashita, A, Endo, M, Higashi, T, Nakatsu, T, Yamada, Y, Oda, J, Kato, H

    Biochemistry   42 ( 19 )   2003

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    DOI: 10.1021/bi0272712

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

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

    Current Biology   13 ( 17 )   2003

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    DOI: 10.1016/S0960-9822(03)00559-1

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

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

    Nature   424 ( 6944 )   2003

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    DOI: 10.1038/nature01780

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

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

    Biophysical Journal   83 ( 5 )   2002

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  • Structure of tropinone reductase-II complexed with NADP(+) and pseudotropine at 1.9 angstrom resolution: Implication for stereospecific substrate binding and catalysis Reviewed

    Yamashita, A, Kato, H, Wakatsuki, S, Tomizaki, T, Nakatsu, T, Nakajima, K, Hashimoto, T, Yamada, Y, Oda, J

    Biochemistry   38 ( 24 )   1999

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    Authorship:Lead author   Publishing type:Research paper (scientific journal)  

    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 )   1998

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    Publishing type:Research paper (scientific journal)  

    DOI: 10.1073/pnas.95.9.4876

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

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

    Acta Crystallographica Section D-Biological Crystallography   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 )   1996

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Books

  • 膜タンパク質工学ハンドブック

    津本, 浩平, 浜窪, 隆雄( Role: Contributor)

    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

    CiNii Books

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  • 口・鼻・耳の感覚メカニズムと応用技術(共著)

    S&T出版  2018  ( ISBN:9784907002701

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  • 膜タンパク質構造研究

    化学同人  2013  ( ISBN:9784759815610

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  • Structural Biology of Membrane Proteins

    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(共著)

    化学同人  2012  ( ISBN:4759815015

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MISC

  • 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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    Authorship:Lead author   Publishing type:Article, review, commentary, editorial, etc. (trade magazine, newspaper, online media)  

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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   86 - 89   2021.3

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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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    Language:English   Publisher:(一社)日本生物物理学会  

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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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    Authorship:Lead author, Corresponding author   Publishing type:Rapid communication, short report, research note, etc. (bulletin of university, research institution)  

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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

  • 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

  • 膜タンパク質構造解析の新展開

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

  • 結晶構造解析における「静止画」から「動画」への試み

    山下 敦子

    日本薬理学雑誌   141 ( 5 )   235 - 239   2013

  • 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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    Language:Japanese   Publisher:The Japanese Association for the Study of Taste and Smell  

    CiNii Article

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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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    Language:English   Publishing type:Research paper, summary (international conference)   Publisher:JAPANESE PHARMACOLOGICAL SOC  

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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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  • 膜タンパク質結晶構造解析と創薬への可能性—神経伝達物質輸送体ホモログLeuTを例にー

    山下敦子

    日本結晶学会誌   52   76 - 80   2010

  • Inhibition mechanism on neurotransmitter transporter homolog revealed by crystallographic analyses

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

    Journal of Pharmacological Sciences   2010

  • 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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    Language:Japanese  

    J-GLOBAL

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

    Atsuko Yamashita

    Seikagaku   2005

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Awards

  • 文部科学大臣表彰 若手科学者賞

    2009  

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