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- EMDB-4160: Imidazoleglycerol-phosphate dehydratase from Saccharomyces cerevisiae -

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Basic information

Entry
Database: EMDB / ID: EMD-4160
TitleImidazoleglycerol-phosphate dehydratase from Saccharomyces cerevisiae
Map dataImidazoleglycerol-phosphate dehydratase
Sample
  • Complex: Imidazoleglycerol-phosphate dehydratase (IGPD) in complex with triazole-phosphonate inhibitor (C384)
    • Protein or peptide: Imidazoleglycerol-phosphate dehydratase
  • Ligand: MANGANESE (II) ION
  • Ligand: [(2R)-2-hydroxy-3-(1H-1,2,4-triazol-1-yl)propyl]phosphonic acid
Function / homology
Function and homology information


imidazoleglycerol-phosphate dehydratase / imidazoleglycerol-phosphate dehydratase activity / histidine biosynthetic process / metal ion binding
Similarity search - Function
Imidazoleglycerol-phosphate dehydratase signature 1. / Imidazoleglycerol-phosphate dehydratase / Imidazoleglycerol-phosphate dehydratase, conserved site / Imidazole glycerol phosphate dehydratase domain superfamily / Imidazoleglycerol-phosphate dehydratase / Imidazoleglycerol-phosphate dehydratase signature 2. / Ribosomal protein S5 domain 2-type fold
Similarity search - Domain/homology
Imidazoleglycerol-phosphate dehydratase
Similarity search - Component
Biological speciesSaccharomyces cerevisiae (strain ATCC 204508 / S288c) (yeast)
Methodsingle particle reconstruction / cryo EM / Resolution: 3.2 Å
AuthorsRawson S / Bisson C / Hurdiss DL / Muench SP
Funding support United Kingdom, 3 items
OrganizationGrant numberCountry
Wellcome Trust(009752/Z/12/Z) and (102572/B/13/Z) United Kingdom
Medical Research Council (United Kingdom)G100567 United Kingdom
Biotechnology and Biological Sciences Research CouncilBB/I003703/1 United Kingdom
CitationJournal: Proc Natl Acad Sci U S A / Year: 2018
Title: Elucidating the structural basis for differing enzyme inhibitor potency by cryo-EM.
Authors: Shaun Rawson / Claudine Bisson / Daniel L Hurdiss / Asif Fazal / Martin J McPhillie / Svetlana E Sedelnikova / Patrick J Baker / David W Rice / Stephen P Muench /
Abstract: Histidine biosynthesis is an essential process in plants and microorganisms, making it an attractive target for the development of herbicides and antibacterial agents. Imidazoleglycerol-phosphate ...Histidine biosynthesis is an essential process in plants and microorganisms, making it an attractive target for the development of herbicides and antibacterial agents. Imidazoleglycerol-phosphate dehydratase (IGPD), a key enzyme within this pathway, has been biochemically characterized in both (IGPD) and (IGPD). The plant enzyme, having been the focus of in-depth structural analysis as part of an inhibitor development program, has revealed details about the reaction mechanism of IGPD, whereas the yeast enzyme has proven intractable to crystallography studies. The structure-activity relationship of potent triazole-phosphonate inhibitors of IGPD has been determined in both homologs, revealing that the lead inhibitor (C348) is an order of magnitude more potent against IGPD than IGPD; however, the molecular basis of this difference has not been established. Here we have used single-particle electron microscopy (EM) to study structural differences between the and IGPD homologs, which could influence the difference in inhibitor potency. The resulting EM maps at ∼3 Å are sufficient to de novo build the protein structure and identify the inhibitor binding site, which has been validated against the crystal structure of the IGPD/C348 complex. The structure of _IGPD reveals that a 24-amino acid insertion forms an extended loop region on the enzyme surface that lies adjacent to the active site, forming interactions with the substrate/inhibitor binding loop that may influence inhibitor potency. Overall, this study provides insights into the IGPD family and demonstrates the power of using an EM approach to study inhibitor binding.
History
DepositionNov 15, 2017-
Header (metadata) releaseNov 29, 2017-
Map releaseFeb 7, 2018-
UpdateDec 11, 2019-
Current statusDec 11, 2019Processing site: PDBe / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 0.116
  • Imaged by UCSF Chimera
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  • Surface view colored by radius
  • Surface level: 0.116
  • Imaged by UCSF Chimera
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  • Surface view with fitted model
  • Atomic models: PDB-6ezm
  • Surface level: 0.116
  • Imaged by UCSF Chimera
  • Download
Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

emd_4160.png

Downloads & links

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Map

FileDownload / File: emd_4160.map.gz / Format: CCP4 / Size: 22.2 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationImidazoleglycerol-phosphate dehydratase
Voxel sizeX=Y=Z: 1.065 Å
Density
Contour LevelBy AUTHOR: 0.116 / Movie #1: 0.116
Minimum - Maximum-0.35065654 - 0.7537618
Average (Standard dev.)0.004652346 (±0.047663864)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions180180180
Spacing180180180
CellA=B=C: 191.70001 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z1.0651.0651.065
M x/y/z180180180
origin x/y/z0.0000.0000.000
length x/y/z191.700191.700191.700
α/β/γ90.00090.00090.000
start NX/NY/NZ-128-128-128
NX/NY/NZ256256256
MAP C/R/S123
start NC/NR/NS000
NC/NR/NS180180180
D min/max/mean-0.3510.7540.005

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Supplemental data

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Sample components

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Entire : Imidazoleglycerol-phosphate dehydratase (IGPD) in complex with tr...

EntireName: Imidazoleglycerol-phosphate dehydratase (IGPD) in complex with triazole-phosphonate inhibitor (C384)
Components
  • Complex: Imidazoleglycerol-phosphate dehydratase (IGPD) in complex with triazole-phosphonate inhibitor (C384)
    • Protein or peptide: Imidazoleglycerol-phosphate dehydratase
  • Ligand: MANGANESE (II) ION
  • Ligand: [(2R)-2-hydroxy-3-(1H-1,2,4-triazol-1-yl)propyl]phosphonic acid

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Supramolecule #1: Imidazoleglycerol-phosphate dehydratase (IGPD) in complex with tr...

SupramoleculeName: Imidazoleglycerol-phosphate dehydratase (IGPD) in complex with triazole-phosphonate inhibitor (C384)
type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1
Source (natural)Organism: Saccharomyces cerevisiae (strain ATCC 204508 / S288c) (yeast)
Recombinant expressionOrganism: Escherichia coli (E. coli)
Molecular weightExperimental: 570 KDa

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Macromolecule #1: Imidazoleglycerol-phosphate dehydratase

MacromoleculeName: Imidazoleglycerol-phosphate dehydratase / type: protein_or_peptide / ID: 1 / Number of copies: 24 / Enantiomer: LEVO / EC number: imidazoleglycerol-phosphate dehydratase
Source (natural)Organism: Saccharomyces cerevisiae (strain ATCC 204508 / S288c) (yeast)
Strain: ATCC 204508 / S288c
Molecular weightTheoretical: 23.867197 KDa
Recombinant expressionOrganism: Escherichia coli (E. coli)
SequenceString: MTEQKALVKR ITNETKIQIA ISLKGGPLAI EHSIFPEKEA EAVAEQATQS QVINVHTGIG FLDHMIHALA KHSGWSLIVE CIGDLHIDD HHTTEDCGIA LGQAFKEALG AVRGVKRFGS GFAPLDEALS RAVVDLSNRP YAVVELGLQR EKVGDLSCEM I PHFLESFA ...String:
MTEQKALVKR ITNETKIQIA ISLKGGPLAI EHSIFPEKEA EAVAEQATQS QVINVHTGIG FLDHMIHALA KHSGWSLIVE CIGDLHIDD HHTTEDCGIA LGQAFKEALG AVRGVKRFGS GFAPLDEALS RAVVDLSNRP YAVVELGLQR EKVGDLSCEM I PHFLESFA EASRITLHVD CLRGKNDHHR SESAFKALAV AIREATSPNG TNDVPSTKGV LM

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Macromolecule #2: MANGANESE (II) ION

MacromoleculeName: MANGANESE (II) ION / type: ligand / ID: 2 / Number of copies: 48 / Formula: MN
Molecular weightTheoretical: 54.938 Da

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Macromolecule #3: [(2R)-2-hydroxy-3-(1H-1,2,4-triazol-1-yl)propyl]phosphonic acid

MacromoleculeName: [(2R)-2-hydroxy-3-(1H-1,2,4-triazol-1-yl)propyl]phosphonic acid
type: ligand / ID: 3 / Number of copies: 24 / Formula: 5LD
Molecular weightTheoretical: 207.124 Da
Chemical component information

ChemComp-5LD:
[(2R)-2-hydroxy-3-(1H-1,2,4-triazol-1-yl)propyl]phosphonic acid

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Experimental details

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Structure determination

Methodcryo EM
Processingsingle particle reconstruction
Aggregation stateparticle

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Sample preparation

Concentration0.5 mg/mL
BufferpH: 7.5
GridModel: Quantifoil R2/2 / Material: COPPER / Pretreatment - Type: GLOW DISCHARGE
VitrificationCryogen name: ETHANE / Chamber humidity: 100 % / Instrument: FEI VITROBOT MARK IV

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELDBright-field microscopy
Sample stageSpecimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Cooling holder cryogen: NITROGEN
Image recordingFilm or detector model: FEI FALCON III (4k x 4k) / Detector mode: INTEGRATING / Number grids imaged: 1 / Number real images: 2827 / Average electron dose: 1.3 e/Å2
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Particle selectionNumber selected: 365498
CTF correctionSoftware - Name: Gctf
Startup modelType of model: EMDB MAP
EMDB ID:

3999


Details: Lowpass filtered to 40 angstroms and rescaled.
Initial angle assignmentType: OTHER
Final angle assignmentType: OTHER
Final reconstructionNumber classes used: 1 / Applied symmetry - Point group: O (octahedral) / Resolution.type: BY AUTHOR / Resolution: 3.2 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: RELION / Details: Relion2.1 auto-refine / Number images used: 11560

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