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- EMDB-18305: Medicago truncatula HISN5 (IGPD) in complex with MN and IG2 -

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Basic information

Entry
Database: EMDB / ID: EMD-18305
TitleMedicago truncatula HISN5 (IGPD) in complex with MN and IG2
Map data
Sample
  • Complex: Complex of imidazole glycerol phosphate dehydratase (HISN5) with inhibitor 2S,3S-IGP (IG2)
    • Protein or peptide: Imidazoleglycerol-phosphate dehydratase
  • Ligand: MANGANESE (II) ION
  • Ligand: (2S,3S)-2,3-dihydroxy-3-(1H-imidazol-5-yl)propyl dihydrogen phosphate
  • Ligand: water
KeywordsMedicago truncatula HISN5 IGPD histidine / LYASE
Function / homology
Function and homology information


imidazoleglycerol-phosphate dehydratase / imidazoleglycerol-phosphate dehydratase activity / L-histidine biosynthetic process
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 speciesMedicago truncatula (barrel medic)
Methodsingle particle reconstruction / cryo EM / Resolution: 2.23 Å
AuthorsWitek W / Ruszkowski M
Funding support Poland, 1 items
OrganizationGrant numberCountry
Polish National Science CentreSonata 2018/31/D/NZ1/03630 Poland
CitationJournal: Front Plant Sci / Year: 2024
Title: Targeting imidazole-glycerol phosphate dehydratase in plants: novel approach for structural and functional studies, and inhibitor blueprinting.
Authors: Wojciech Witek / Joanna Sliwiak / Michal Rawski / Milosz Ruszkowski /
Abstract: The histidine biosynthetic pathway (HBP) is targeted for herbicide design with preliminary success only regarding imidazole-glycerol phosphate dehydratase (IGPD, EC 4.2.1.19), or HISN5, as referred ...The histidine biosynthetic pathway (HBP) is targeted for herbicide design with preliminary success only regarding imidazole-glycerol phosphate dehydratase (IGPD, EC 4.2.1.19), or HISN5, as referred to in plants. HISN5 catalyzes the sixth step of the HBP, in which imidazole-glycerol phosphate (IGP) is dehydrated to imidazole-acetol phosphate. In this work, we present high-resolution cryoEM and crystal structures of HISN5 (HISN5) in complexes with an inactive IGP diastereoisomer and with various other ligands. HISN5 can serve as a new model for plant HISN5 structural studies, as it enables resolving protein-ligand interactions at high (2.2 Å) resolution using cryoEM. We identified ligand-binding hotspots and characterized the features of plant HISN5 enzymes in the context of the HISN5-targeted inhibitor design. Virtual screening performed against millions of small molecules not only revealed candidate molecules but also identified linkers for fragments that were experimentally confirmed to bind. Based on experimental and computational approaches, this study provides guidelines for designing symmetric HISN5 inhibitors that can reach two neighboring active sites. Finally, we conducted analyses of sequence similarity networks revealing that plant HISN5 enzymes derive from cyanobacteria. We also adopted a new approach to measure HISN5 enzymatic activity using isothermal titration calorimetry and enzymatically synthesized IGP.
History
DepositionAug 23, 2023-
Header (metadata) releaseApr 10, 2024-
Map releaseApr 10, 2024-
UpdateApr 10, 2024-
Current statusApr 10, 2024Processing site: PDBe / Status: Released

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Structure visualization

Supplemental images

emd_18305.png

Downloads & links

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Map

FileDownload / File: emd_18305.map.gz / Format: CCP4 / Size: 103 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
0.86 Å/pix.
x 300 pix.
= 258. Å		0.86 Å/pix.
x 300 pix.
= 258. Å		0.86 Å/pix.
x 300 pix.
= 258. Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 0.86 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.743
Minimum - Maximum-3.2042494 - 6.1224284
Average (Standard dev.)-0.0009380566 (±0.23926952)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions300300300
Spacing300300300
CellA=B=C: 258.0 Å
α=β=γ: 90.0 °

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Supplemental data

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Half map: #2

Fileemd_18305_half_map_1.map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: #1

Fileemd_18305_half_map_2.map
Projections & Slices
AxesZYX

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Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Sample components

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Entire : Complex of imidazole glycerol phosphate dehydratase (HISN5) with ...

EntireName: Complex of imidazole glycerol phosphate dehydratase (HISN5) with inhibitor 2S,3S-IGP (IG2)
Components
  • Complex: Complex of imidazole glycerol phosphate dehydratase (HISN5) with inhibitor 2S,3S-IGP (IG2)
    • Protein or peptide: Imidazoleglycerol-phosphate dehydratase
  • Ligand: MANGANESE (II) ION
  • Ligand: (2S,3S)-2,3-dihydroxy-3-(1H-imidazol-5-yl)propyl dihydrogen phosphate
  • Ligand: water

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Supramolecule #1: Complex of imidazole glycerol phosphate dehydratase (HISN5) with ...

SupramoleculeName: Complex of imidazole glycerol phosphate dehydratase (HISN5) with inhibitor 2S,3S-IGP (IG2)
type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1
Source (natural)Organism: Medicago truncatula (barrel medic)
Molecular weightTheoretical: 22.5 kDa/nm

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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: Medicago truncatula (barrel medic)
Molecular weightTheoretical: 22.571293 KDa
Recombinant expressionOrganism: Escherichia coli (E. coli)
SequenceString: PTFPIDSGAR IGEMKRVTKE TNVSVKINLD GTGVADNSSG IPFLDHMLDQ LASHGLFDVH VKATGDTHID DHHTNEDVAL AIGTALLQA LGDRKGINRF GNFSAPLDEA LVHVSLDLSG RPHLGYDLNI PTQRVGKYDT QLVEHFFQSL VNTSGMTLHI R QFSGTNSH ...String:
PTFPIDSGAR IGEMKRVTKE TNVSVKINLD GTGVADNSSG IPFLDHMLDQ LASHGLFDVH VKATGDTHID DHHTNEDVAL AIGTALLQA LGDRKGINRF GNFSAPLDEA LVHVSLDLSG RPHLGYDLNI PTQRVGKYDT QLVEHFFQSL VNTSGMTLHI R QFSGTNSH HIIEATFKAF ARALRQATEY DTRRRGTIPS SKGVLSRS

UniProtKB: Imidazoleglycerol-phosphate dehydratase

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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: (2S,3S)-2,3-dihydroxy-3-(1H-imidazol-5-yl)propyl dihydrogen phosphate

MacromoleculeName: (2S,3S)-2,3-dihydroxy-3-(1H-imidazol-5-yl)propyl dihydrogen phosphate
type: ligand / ID: 3 / Number of copies: 24 / Formula: IG2
Molecular weightTheoretical: 238.135 Da
Chemical component information

ChemComp-IG2:
(2S,3S)-2,3-dihydroxy-3-(1H-imidazol-5-yl)propyl dihydrogen phosphate

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Macromolecule #4: water

MacromoleculeName: water / type: ligand / ID: 4 / Number of copies: 1131 / Formula: HOH
Molecular weightTheoretical: 18.015 Da
Chemical component information

ChemComp-HOH:
WATER

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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

Concentration1 mg/mL
BufferpH: 8
GridModel: Quantifoil / Material: COPPER / Mesh: 300 / Support film - Material: CARBON / Support film - topology: HOLEY / Pretreatment - Type: GLOW DISCHARGE
VitrificationCryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 277 K / Instrument: FEI VITROBOT MARK IV

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Image recordingFilm or detector model: FEI FALCON III (4k x 4k) / Detector mode: COUNTING / Number grids imaged: 1 / Number real images: 3330 / Average electron dose: 40.0 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: SPOT SCAN / Imaging mode: BRIGHT FIELD / Cs: 2.7 mm / Nominal defocus max: 3.5 µm / Nominal defocus min: 1.0 µm
Sample stageSpecimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Cooling holder cryogen: NITROGEN
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Startup modelType of model: NONE
Final reconstructionNumber classes used: 14 / Applied symmetry - Point group: O (octahedral) / Resolution.type: BY AUTHOR / Resolution: 2.23 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: cryoSPARC (ver. 3.3.2) / Number images used: 605614
Initial angle assignmentType: MAXIMUM LIKELIHOOD
Final angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: cryoSPARC
Final 3D classificationNumber classes: 50 / Software - Name: cryoSPARC

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Atomic model buiding 1

Initial modelPDB ID:

7oj5


Chain - Source name: PDB / Chain - Initial model type: experimental model
RefinementSpace: REAL / Protocol: RIGID BODY FIT
Output model

PDB-8qav:
Medicago truncatula HISN5 (IGPD) in complex with MN and IG2

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