KINDAI UNIVERSITY


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SHIMAMOTO Shigeru

Profile

FacultyDepartment of Life Science / Graduate School of Science and Engineering Research
PositionLecturer
Degree
Commentator Guidehttps://www.kindai.ac.jp/meikan/906-shimamoto-shigeru.html
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Last Updated :2020/07/08

Education and Career

Academic & Professional Experience

  •   2015 04 ,  - 現在, Faculty of Science and Engineering, Department of Life Science, Kindai University
  •   2011 04 ,  - 2014 03 , Faculty of Science and Engineering, Department of Life Science, Kindai University

Research Activities

Research Areas

  • Life sciences, Structural biochemistry
  • Nanotechnology/Materials, Molecular biochemistry
  • Life sciences, Pharmaceuticals - analytical and physicochemistry

Research Interests

  • protein folding, ITC, NMR

Published Papers

  • N-terminal HCV core protein fragment decreases 20S proteasome activity in the presence of PA28γ., Zheng Y, Shimamoto S, Maruno T, Kobayashi Y, Matsuura Y, Kawahara K, Yoshida T, Ohkubo T, Biochemical and biophysical research communications, Biochemical and biophysical research communications, 509(2), 590 - 595, Feb. 2019 , Refereed
  • H-1, C-13, and N-15 resonance assignments of mouse lipocalin-type prostaglandin D synthase/substrate analog complex, Shigeru Shimamoto, Hiroko Maruo, Takuya Yoshida, Tadayasu Ohkubo, BIOMOLECULAR NMR ASSIGNMENTS, BIOMOLECULAR NMR ASSIGNMENTS, 8(1), 129 - 132, Apr. 2014 , Refereed
    Summary:Lipocalin-type Prostaglandin D synthase (L-PGDS) acts as the PGD(2)-synthesizing enzyme in the brain of various mammalian species. It belongs to the lipocalin superfamily and is the first member of this family to be recognized as an enzyme. Although the solution and crystal structure of L-PGDS has been determined to understand the molecular mechanism of catalytic reaction, the structural analysis of L-PGDS in complex with its substrate remains to be performed. Here, we present the nearly complete assignment of the backbone and side chain resonances of L-PGDS/substrate analog (U-46619) complex. This study lays the essential basis for further understanding the substrate recognition mechanism of L-PGDS.
  • Chemical methods for producing disulfide bonds in peptides and proteins to study folding regulation., Okumura M, Shimamoto S, Hidaka Y, Current protocols in protein science, Current protocols in protein science, 76, 28.7.1 - 13, Apr. 2014 , Refereed
  • Chemical methods and approaches to the regioselective formation of multiple disulfide bonds., Shimamoto S, Katayama H, Okumura M, Hidaka Y, Curr. Protoc. Protein Sci., Curr. Protoc. Protein Sci., 76, 28.8.1 - 28.8.28, Apr. 2014 , Refereed
  • Folding of Peptides and Proteins: Role of Disulfide Bonds, Recent Developments., Hidaka Y, Shimamoto S, BioMol. Concepts, BioMol. Concepts, 4(6), 597 - 604, Dec. 2013 , Refereed
  • Crystal structure of the dog allergen Can f 6 and structure-based implications of its cross-reactivity with the cat allergen Fel d 4., Kenji Yamamoto, Osamu Ishibashi, Keisuke Sugiura, Miki Ubatani, Masaya Sakaguchi, Masatoshi Nakatsuji, Shigeru Shimamoto, Masanori Noda, Susumu Uchiyama, Yuma Fukutomi, Shigenori Nishimura, Takashi Inui, Scientific reports, Scientific reports, 9(1), 1503 - 1503, Feb. 06 2019 , Refereed
    Summary:Several dog allergens cause allergic reactions in humans worldwide. Seven distinct dog allergens, designated Canis familiaris allergen 1 to 7 (Can f 1-Can f 7), have been identified thus far. Can f 6 shows high sequence similarity and cross-reactivity with Fel d 4 and Equ c 1, major cat and horse allergens, respectively. This study was conducted on the allergenic epitopes of Can f 6 based on its structural characterization. We demonstrated that sera from 18 out of 38 (47%) dog-sensitized patients reacted to recombinant Can f 6 protein (rCan f 6). We then determined the crystal structure of rCan f 6 by X-ray crystallography, which exhibited a conserved tertiary structural architecture found in lipocalin family proteins. Based on the tertiary structure and sequence similarities with Fel d 4 and Equ c 1, we predicted three IgE-recognizing sites that are possibly involved in cross-reactivity. Substituting three successive amino acids in these sites to triple alanine decreased IgE reactivity to the allergen. However, the degree of reduction in IgE reactivity largely depended on the site mutated and the serum used, suggesting that Can f 6 is a polyvalent allergen containing multiple epitopes and Can f 6-reactive sera contain varied amounts of IgE recognising individual Can f 6 epitopes including those predicted in this study. We also demonstrated that the predicted epitopes are partly involved in IgE cross-reactivity to Fel d 4. Interestingly, the effect of the mutation depended on whether the protein was structured or denatured, indicating that the bona fide tertiary structure of Can f 6 is essential in determining its IgE epitopes.
  • Thermodynamic and NMR analyses of NADPH binding to lipocalin-type prostaglandin D synthase, Shubin Qin, Shigeru Shimamoto, Takahiro Maruno, Yuji Kobayashi, Kazuki Kawahara, Takuya Yoshida, Tadayasu Ohkubo, BIOCHEMICAL AND BIOPHYSICAL RESEARCH COMMUNICATIONS, BIOCHEMICAL AND BIOPHYSICAL RESEARCH COMMUNICATIONS, 468(1-2), 234 - 239, Dec. 2015 , Refereed
    Summary:Lipocalin-type prostaglandin D synthase (L-PGDS) is one of the most abundant proteins in human cerebrospinal fluid (CSF) with dual functions as a prostaglandin D-2 (PGD(2)) synthase and a transporter of lipophilic ligands. Recent studies revealed that L-PGDS plays important roles in protecting against various neuronal diseases induced by reactive oxygen species (ROS). However, the molecular mechanisms of such protective actions of L-PGDS remain unknown. In this study, we conducted thermodynamic and nuclear magnetic resonance (NMR) analyses, and demonstrated that L-PGDS binds to nicotinamide coenzymes, including NADPH, NADP(+), and NADH. Although a hydrophilic ligand is not common for L-PGDS, these ligands, especially NADPH showed specific interaction with L-PGDS at the upper pocket of its ligand-binding cavity with an unusually bifurcated shape. The binding affinity of L-PGDS for NADPH was comparable to that previously reported for NADPH oxidases and NADPH in vitro. These results suggested that L-PGDS potentially attenuates the activities of NADPH oxidases through interaction with NADPH. Given that NADPH is the substrate for NADPH oxidases that play key roles in neuronal cell death by generating excessive ROS, these results imply a novel linkage between L-PGDS and ROS. (C) 2015 Elsevier Inc. All rights reserved.
  • Effects of positively charged redox molecules on disulfide-coupled protein folding, Masaki Okumura, Shigeru Shimamoto, Takeyoshi Nakanishi, Yu-ichiro Yoshida, Tadafumi Konogami, Shogo Maeda, Yuji Hidaka, FEBS LETTERS, FEBS LETTERS, 586(21), 3926 - 3930, Nov. 2012 , Refereed
    Summary:In vitro folding of disulfide-containing proteins is generally regulated by redox molecules, such as glutathione. However, the role of the cross-disulfide-linked species formed between the redox molecule and the protein as a folding intermediate in the folding mechanism is poorly understood. In the present study, we investigated the effect of the charge on a redox molecule on disulfide-coupled protein folding. Several types of aliphatic thiol compounds including glutathione were examined for the folding of disulfide-containing-proteins, such as lysozyme and prouroguanylin. The results indicate that the positive charge and its dispersion play a critical role in accelerating disulfide-coupled protein folding. (C) 2012 Federation of European Biochemical Societies. Published by Elsevier B.V. All rights reserved.
  • A chemical method for investigating disulfide-coupled peptide and protein folding, Masaki Okumura, Shigeru Shimamoto, Yuji Hidaka, FEBS JOURNAL, FEBS JOURNAL, 279(13), 2283 - 2295, Jul. 2012 , Refereed
    Summary:Investigations of protein folding have largely involved studies using disulfide-containing proteins, as disulfide-coupled folding of proteins permits the folding intermediates to be trapped and their conformations determined. Over the last decade, a combination of new biotechnical and chemical methodology has resulted in a remarkable acceleration in our understanding of the mechanism of disulfide-coupled protein folding. In particular, expressed protein ligation, a combination of native chemical ligation and an intein-based approach, permits specifically labeled proteins to be easily produced for studies of protein folding using biophysical methods, such as NMR spectroscopy and X-ray crystallography. A method for regio-selective formation of disulfide bonds using chemical procedures has also been established. This strategy is particularly relevant for the study of disulfide-coupled protein folding, and provides us not only with the native conformation, but also the kinetically trapped topological isomer with native disulfide bonds. Here we review recent developments and applications of biotechnical and chemical methods to investigations of disulfide-coupled peptide and protein folding. Chemical additives designed to accelerate correct protein folding and to avoid non-specific aggregation are also discussed.
  • Drug delivery system for poorly water-soluble compounds using lipocalin-type prostaglandin D synthase, Ayano Fukuhara, Hidemitsu Nakajima, Yuya Miyamoto, Katsuaki Inoue, Satoshi Kume, Young-Ho Lee, Masanori Noda, Susumu Uchiyama, Shigeru Shimamoto, Shigenori Nishimura, Tadayasu Ohkubo, Yuji Goto, Tadayoshi Takeuchi, Takashi Inui, JOURNAL OF CONTROLLED RELEASE, JOURNAL OF CONTROLLED RELEASE, 159(1), 143 - 150, Apr. 2012 , Refereed
    Summary:Lipocalin-type prostaglandin D synthase (L-PGDS) is a member of the lipocalin superfamily and a secretory lipid-transporter protein, which binds a wide variety of hydrophobic small molecules. Here we show the feasibility of a novel drug delivery system (DDS), utilizing L-PGDS, for poorly water-soluble compounds such as diazepam (DZP), a major benzodiazepine anxiolytic drug, and 6-nitro-7-sulfamoylbenzo[f] quinoxaline-2,3-dione (NBQX), an a-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) receptor antagonist and anticonvulsant. Calorimetric experiments revealed for both compounds that each L-PGDS held three molecules with high binding affinities. By mass spectrometry, the 1:3 complex of L-PGDS and NBQX was observed. L-PGDS of 500 mu M increased the solubility of DZP and NBQX 7- and 2-fold, respectively, compared to PBS alone. To validate the potential of L-PGDS as a drug delivery vehicle in vivo, we have proved the prospective effects of these compounds via two separate delivery strategies. First, the oral administration of a DZP/L-PGDS complex in mice revealed an increased duration of pentobarbital-induced loss of righting reflex. Second, the intravenous treatment of ischemic gerbils with NBQX/L-PGDS complex showed a protective effect on delayed neuronal cell death at the hippocampal CA1 region. We propose that our novel DDS could facilitate pharmaceutical development and clinical usage of various water-insoluble compounds. (C) 2011 Elsevier B.V. All rights reserved.
  • The Middle Region of an HP1-binding Protein, HP1-BP74, Associates with Linker DNA at the Entry/Exit Site of Nucleosomal DNA, Kayoko Hayashihara, Susumu Uchiyama, Shigeru Shimamoto, Shouhei Kobayashi, Miroslav Tomschik, Hidekazu Wakamatsu, Daisuke No, Hiroki Sugahara, Naoto Hori, Masanori Noda, Tadayasu Ohkubo, Jordanka Zlatanova, Sachihiro Matsunaga, Kiichi Fukui, JOURNAL OF BIOLOGICAL CHEMISTRY, JOURNAL OF BIOLOGICAL CHEMISTRY, 285(9), 6498 - 6507, Feb. 2010 , Refereed
    Summary:In higher eukaryotic cells, DNA molecules are present as chromatin fibers, complexes of DNA with various types of proteins; chromatin fibers are highly condensed in metaphase chromosomes during mitosis. Although the formation of the metaphase chromosome structure is essential for the equal segregation of replicated chromosomal DNA into the daughter cells, the mechanism involved in the organization of metaphase chromosomes is poorly understood. To identify proteins involved in the formation and/or maintenance of metaphase chromosomes, we examined proteins that dissociated from isolated human metaphase chromosomes by 0.4 M NaCl treatment; this treatment led to significant chromosome decondensation, but the structure retained the core histones. One of the proteins identified, HP1-BP74 (heterochromatin protein 1-binding protein 74), composed of 553 amino acid residues, was further characterized. HP1-BP74 middle region (BP74Md), composed of 178 amino acid residues (Lys(97)-Lys(274)), formed a chromatosome-like structure with reconstituted mononucleosomes and protected the linker DNA from micrococcal nuclease digestion by similar to 25 bp. The solution structure determined by NMR revealed that the globular domain (Met(153)-Thr(237)) located within BP74Md possesses a structure similar to that of the globular domain of linker histones, which underlies its nucleosome binding properties. Moreover, we confirmed that BP74Md and full-length HP1-BP74 directly binds to HP1 (heterochromatin protein 1) and identified the exact sites responsible for this interaction. Thus, we discovered that HP1-BP74 directly binds to HP1, and its middle region associates with linker DNA at the entry/exit site of nucleosomal DNA in vitro.
  • Structural analysis of lipocalin-type prostaglandin D synthase complexed with biliverdin by small-angle X-ray scattering and multi-dimensional NMR, Yuya Miyamoto, Shigenori Nishimura, Katsuaki Inoue, Shigeru Shimamoto, Takuya Yoshida, Ayano Fukuhara, Mao Yamada, Yoshihiro Urade, Naoto Yagi, Tadayasu Ohkubo, Takashi Inui, JOURNAL OF STRUCTURAL BIOLOGY, JOURNAL OF STRUCTURAL BIOLOGY, 169(2), 209 - 218, Feb. 2010 , Refereed
    Summary:Lipocalin-type prostaglandin D synthase (L-PGDS) acts as both a PGD(2) synthase and an extracellular transporter for small lipophilic molecules. From a series of biochemical studies, it has been found that L-PGDS has an ability to bind a variety of lipophilic ligands such as biliverdin, bilirubin and retinoids in vitro. Therefore, we considered that it is necessary to clarify the molecular structure of L-PGDS upon binding ligand in order to understand the physiological relevance of L-PGDS as a transporter protein. We investigated a molecular structure of L-PGDS/biliverdin complex by small-angle X-ray scattering (SAXS) and multi-dimensional NMR measurements, and characterized the binding mechanism in detail. SAXS measurements revealed that L-PGDS has a globular shape and becomes compact by 1.3 angstrom in radius of gyration on binding biliverdin. NMR experiments revealed that L-PGDS possessed an eight-stranded antiparallel beta-barrel forming a central cavity. Upon the titration with biliverdin, some cross-peaks for residues surrounding the cavity and EF-loop and H2-helix above the beta-barrel shifted, and the intensity of other cross-peaks decreased with signal broadenings in (1)H-(15)N heteronuclear single quantum coherence spectra. These results demonstrate that L-PGDS holds biliverdin within the beta-barrel, and the conformation of the loop regions above the beta-barrel changes upon binding biliverdin. Through such a conformational change, the whole molecule of L-PGDS becomes compact. (C) 2009 Elsevier Inc. All rights reserved.
  • Biochemical, Functional, and Pharmacological Characterization of AT-56, an Orally Active and Selective Inhibitor of Lipocalin-type Prostaglandin D Synthase, Daisuke Irikura, Kosuke Aritake, Nanae Nagata, Toshihiko Maruyama, Shigeru Shimamoto, Yoshihiro Urade, JOURNAL OF BIOLOGICAL CHEMISTRY, JOURNAL OF BIOLOGICAL CHEMISTRY, 284(12), 7623 - 7630, Mar. 2009 , Refereed
    Summary:We report here that 4-dibenzo[a,d] cyclohepten-5-ylidene-1[4-(2H-tetrazol-5-yl)-butyl]-piperidine (AT-56) is an orally active and selective inhibitor of lipocalin-type prostaglandin (PG) D synthase (L-PGDS). AT-56 inhibited human and mouse L-PGDSs in a concentration (3-250 mu M)-dependent manner but did not affect the activities of hematopoietic PGD synthase (H-PGDS), cyclooxygenase-1 and -2, and microsomal PGE synthase1. AT-56 inhibited the L-PGDS activity in a competitive manner against the substrate-PGH(2) (K(m) = 14 mu M) with a K(i) value of 75 mu M but did not inhibit the binding of 13-cis-retinoic acid, a nonsubstrate lipophilic ligand, to L-PGDS. NMR titration analysis revealed that AT-56 occupied the catalytic pocket, but not the retinoid-binding pocket, of L-PGDS. AT-56 inhibited the production of PGD(2) by L-PGDS-expressing human TE-671 cells after stimulation with Ca(2+) ionophore (5 mu M A23187) with an IC(50) value of about 3 mu M without affecting their production of PGE(2) and PGF(2 alpha) but had no effect on the PGD2 production by H-PGDS-expressing human megakaryocytes. Orally administered AT-56 (< 30 mg/ kg body weight) decreased the PGD2 production to 40% in the brain of H-PGDS-deficient mice after a stab wound injury in a dose-dependent manner without affecting the production of PGE2 and PGF(2 alpha) and also suppressed the accumulation of eosinophils and monocytes in the bronco-alveolar lavage fluid from the antigen-induced lung inflammation model of human L-PGDS-transgenic mice.
  • NMR solution structure of lipocalin-type prostaglandin D synthase - Evidence for partial overlapping of catalytic pocket and retinoic acid-binding pocket within the central cavity, Shigeru Shimamoto, Takuya Yoshida, Takashi Inui, Keigo Gohda, Yuji Kobayashi, Ko Fujimori, Toshiharu Tsurumura, Kosuke Aritake, Yoshihiro Urade, Tadayasu Ohkubo, JOURNAL OF BIOLOGICAL CHEMISTRY, JOURNAL OF BIOLOGICAL CHEMISTRY, 282(43), 31373 - 31379, Oct. 2007 , Refereed
    Summary:Lipocalin-type prostaglandin (PG) D synthase (L-PGDS) catalyzes the isomerization of PGH(2), a common precursor of various prostanoids, to produce PGD(2), an endogenous somnogen and nociceptive modulator, in the brain. L-PGDS is a member of the lipocalin superfamily and binds lipophilic substances, such as retinoids and bile pigments, suggesting that L-PGDS is a dual functional protein acting as a PGD(2)-synthesizing enzyme and a transporter for lipophilic ligands. In this study we determined by NMR the three-dimensional structure of recombinant mouse L-PGDS with the catalytic residue Cys-65. The structure of L-PGDS exhibited the typical lipocalin fold, consisting of an eight-stranded, antiparallel beta-barrel and a long alpha-helix associated with the outer surface of the barrel. The interior of the barrel formed a hydrophobic cavity opening to the upper end of the barrel, the size of which was larger than those of other lipocalins, and the cavity contained two pockets. Molecular docking studies, based on the result of NMR titration experiments with retinoic acid and PGH2 analog, revealed that PGH(2) almost fully occupied the hydrophilic pocket 1, in which Cys-65 was located and all-trans-retinoic acid occupied the hydrophobic pocket 2, in which amino acid residues important for retinoid binding in other lipocalins were well conserved. Mutational and kinetic studies provide the direct evidence for the PGH(2) binding mode. These results indicated that the two binding sites for PGH2 and retinoic acid in the large cavity of L-PGDS were responsible for the broad ligand specificity of L-PGDS and the non-competitive inhibition of L-PGDS activity by retinoic acid.

Misc

  • ITC Analysis of Ligand Binding to Lipocalin-type Prostaglandin D Synthase, Shigeru Shimamoto, Netsu Sokutei, 44, 3, 108, 116,   2017 07 , Refereed, 招待有り
  • Ligand Recognition Mechanism of Lipocalin-type Prostaglandin D Synthase, SHIMAMOTO Shigeru, YOSHIDA Takuya, OHKUBO Tadayasu, YAKUGAKU ZASSHI, 131, 11, 1575, 1581,   2011 , Refereed, 招待有り, 10.1248/yakushi.131.1575, http://ci.nii.ac.jp/naid/130001491111
    Summary:Lipocalin-type prostaglandin (PG) D synthase (L-PGDS) is a multi functional protein acting as a PGD2 synthesizing enzyme, a transporter or scavenger of various lipophilic ligands, and an amyloid β chaperon in the brain. L-PGDS is a member of the lipocalin superfamily and has the ability to bind various lipophilic molecules such as prostanoid, retinoid, bile pigment, and amyloid β peptide. However, the molecular mechanism for a wide variety of ligand binding has not been well understood. In this study, we determined by NMR the structure of recombinant mouse L-PGDS and L-PGDS/PGH2 analog complex. L-PGDS has the typical lipocalin fold, consisting of an eight-stranded β-barrel and a long α-helix. The interior of the barrel formed a hydrophobic cavity opening to the upper end of the barrel, the size of which was larger than those of other lipocalins and the cavity contained two pockets. Kinetic studies and molecular docking studies based on the result of NMR titration experiments provide the direct evidence for two binding sites for PGH2 and retinoic acid in the large cavity of L-PGDS. Structural comparison of L-PGDS/U-46619 complex with apo-L-PGDS showed that the H2-helix, CD-loop, and EF-loop located at the upper end of the β-barrel change the conformation to cover the entry of the cavity upon U-46619 binding. These results indicated that the two binding sites in the large cavity and induced fit mechanism were responsible for the broad ligand specificity of L-PGDS.