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- EMDB-15084: cryo-EM structure of thioredoxin glutathione reductase in complex... -

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Basic information

Entry
Database: EMDB / ID: EMD-15084
Titlecryo-EM structure of thioredoxin glutathione reductase in complex with a non-competitive inhibitor
Map data
Sample
  • Complex: TGR in complex with an inhibitor
    • Complex: TGR complex with inhibitor
      • Protein or peptide: Thioredoxin glutathione reductase
  • Ligand: FLAVIN-ADENINE DINUCLEOTIDEFlavin adenine dinucleotide
  • Ligand: (2~{R},3~{R},4~{S},5~{R})-2-[3-[[[(1~{R},2~{R},3~{R},5~{S})-2,6,6-trimethyl-3-bicyclo[3.1.1]heptanyl]amino]methyl]indol-1-yl]oxane-3,4,5-triol
Keywordsinhibitor / complex / flavoreductase / FLAVOPROTEIN
Function / homology
Function and homology information


thioredoxin-disulfide reductase / thioredoxin-disulfide reductase (NADPH) activity / cell redox homeostasis / flavin adenine dinucleotide binding / metal ion binding
Similarity search - Function
Thioredoxin/glutathione reductase selenoprotein / Glutaredoxin, eukaryotic/virial / Glutaredoxin active site / Glutaredoxin active site. / : / Glutaredoxin / Glutaredoxin / Glutaredoxin domain profile. / Pyridine nucleotide-disulphide oxidoreductase, class I / Pyridine nucleotide-disulphide oxidoreductase, class I, active site ...Thioredoxin/glutathione reductase selenoprotein / Glutaredoxin, eukaryotic/virial / Glutaredoxin active site / Glutaredoxin active site. / : / Glutaredoxin / Glutaredoxin / Glutaredoxin domain profile. / Pyridine nucleotide-disulphide oxidoreductase, class I / Pyridine nucleotide-disulphide oxidoreductase, class I, active site / Pyridine nucleotide-disulphide oxidoreductases class-I active site. / Pyridine nucleotide-disulphide oxidoreductase, dimerisation domain / Pyridine nucleotide-disulphide oxidoreductase, dimerisation domain / FAD/NAD-linked reductase, dimerisation domain superfamily / FAD/NAD(P)-binding domain / Pyridine nucleotide-disulphide oxidoreductase / FAD/NAD(P)-binding domain superfamily / Thioredoxin-like superfamily
Similarity search - Domain/homology
thioredoxin-disulfide reductase
Similarity search - Component
Biological speciesSchistosoma mansoni (invertebrata)
Methodsingle particle reconstruction / cryo EM / Resolution: 3.6 Å
AuthorsArdini M / Angelucci F / Fata F / Gabriele F / Effantin G / Ling W / Williams DL / Petukhova VZ / Petukhov PA
Funding support United States, 1 items
OrganizationGrant numberCountry
National Institutes of Health/National Institute Of Allergy and Infectious Diseases (NIH/NIAID)R33AI127635 United States
CitationJournal: Nat Commun / Year: 2023
Title: Non-covalent inhibitors of thioredoxin glutathione reductase with schistosomicidal activity in vivo.
Authors: Valentina Z Petukhova / Sammy Y Aboagye / Matteo Ardini / Rachel P Lullo / Francesca Fata / Margaret E Byrne / Federica Gabriele / Lucy M Martin / Luke N M Harding / Vamshikrishna Gone / ...Authors: Valentina Z Petukhova / Sammy Y Aboagye / Matteo Ardini / Rachel P Lullo / Francesca Fata / Margaret E Byrne / Federica Gabriele / Lucy M Martin / Luke N M Harding / Vamshikrishna Gone / Bikash Dangi / Daniel D Lantvit / Dejan Nikolic / Rodolfo Ippoliti / Grégory Effantin / Wai Li Ling / Jeremy J Johnson / Gregory R J Thatcher / Francesco Angelucci / David L Williams / Pavel A Petukhov /
Abstract: Only praziquantel is available for treating schistosomiasis, a disease affecting more than 200 million people. Praziquantel-resistant worms have been selected for in the lab and low cure rates from ...Only praziquantel is available for treating schistosomiasis, a disease affecting more than 200 million people. Praziquantel-resistant worms have been selected for in the lab and low cure rates from mass drug administration programs suggest that resistance is evolving in the field. Thioredoxin glutathione reductase (TGR) is essential for schistosome survival and a validated drug target. TGR inhibitors identified to date are irreversible and/or covalent inhibitors with unacceptable off-target effects. In this work, we identify noncovalent TGR inhibitors with efficacy against schistosome infections in mice, meeting the criteria for lead progression indicated by WHO. Comparisons with previous in vivo studies with praziquantel suggests that these inhibitors outperform the drug of choice for schistosomiasis against juvenile worms.
History
DepositionJun 1, 2022-
Header (metadata) releaseJun 14, 2023-
Map releaseJun 14, 2023-
UpdateJul 5, 2023-
Current statusJul 5, 2023Processing site: PDBe / Status: Released

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Structure visualization

Supplemental images

emd_15084.png

Downloads & links

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Map

FileDownload / File: emd_15084.map.gz / Format: CCP4 / Size: 103 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Voxel sizeX=Y=Z: 1.145 Å
Density
Contour LevelBy AUTHOR: 0.012
Minimum - Maximum-0.01418814 - 0.06508718
Average (Standard dev.)0.000039421706 (±0.00147939)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderYXZ
Origin000
Dimensions300300300
Spacing300300300
CellA=B=C: 343.5 Å
α=β=γ: 90.0 °

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Supplemental data

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Additional map: This is the final sharpened map made by...

Fileemd_15084_additional_1.map
AnnotationThis is the final sharpened map made by Phenix (version 1.19.2-4158) local anisotropic sharpening from the refined fullmap_handcoot_map.ccp4
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: #2

Fileemd_15084_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_15084_half_map_2.map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Sample components

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Entire : TGR in complex with an inhibitor

EntireName: TGR in complex with an inhibitor
Components
  • Complex: TGR in complex with an inhibitor
    • Complex: TGR complex with inhibitor
      • Protein or peptide: Thioredoxin glutathione reductase
  • Ligand: FLAVIN-ADENINE DINUCLEOTIDEFlavin adenine dinucleotide
  • Ligand: (2~{R},3~{R},4~{S},5~{R})-2-[3-[[[(1~{R},2~{R},3~{R},5~{S})-2,6,6-trimethyl-3-bicyclo[3.1.1]heptanyl]amino]methyl]indol-1-yl]oxane-3,4,5-triol

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Supramolecule #1: TGR in complex with an inhibitor

SupramoleculeName: TGR in complex with an inhibitor / type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1
Details: Recombinant TGR bound non-covalently to a synthetic chimeric compound
Source (natural)Organism: Schistosoma mansoni (invertebrata)

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Supramolecule #2: TGR complex with inhibitor

SupramoleculeName: TGR complex with inhibitor / type: complex / ID: 2 / Parent: 1 / Macromolecule list: #1
Source (natural)Organism: Schistosoma mansoni (invertebrata)

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Macromolecule #1: Thioredoxin glutathione reductase

MacromoleculeName: Thioredoxin glutathione reductase / type: protein_or_peptide / ID: 1 / Number of copies: 2 / Enantiomer: LEVO / EC number: ec: 1.6.4.5
Source (natural)Organism: Schistosoma mansoni (invertebrata)
Molecular weightTheoretical: 65.061145 KDa
Recombinant expressionOrganism: Escherichia coli BL21(DE3) (bacteria)
SequenceString: MPPADGTSQW LRKTVDSAAV ILFSKTTCPY CKKVKDVLAE AKIKHATIEL DQLSNGSAIQ KCLASFSKIE TVPQMFVRGK FIGDSQTVL KYYSNDELAG IVNESKYDYD LIVIGGGSGG LAAGKEAAKY GAKTAVLDYV EPTPIGTTWG LGGTCVNVGC I PKKLMHQA ...String:
MPPADGTSQW LRKTVDSAAV ILFSKTTCPY CKKVKDVLAE AKIKHATIEL DQLSNGSAIQ KCLASFSKIE TVPQMFVRGK FIGDSQTVL KYYSNDELAG IVNESKYDYD LIVIGGGSGG LAAGKEAAKY GAKTAVLDYV EPTPIGTTWG LGGTCVNVGC I PKKLMHQA GLLSHALEDA EHFGWSLDRS KISHNWSTMV EGVQSHIGSL NWGYKVALRD NQVTYLNAKG RLISPHEVQI TD KNQKVST ITGNKIILAT GERPKYPEIP GAVEYGITSD DLFSLPYFPG KTLVIGASYV ALECAGFLAS LGGDVTVMVR SIL LRGFDQ QMAEKVGDYM ENHGVKFAKL CVPDEIKQLK VVDTENNKPG LLLVKGHYTD GKKFEEEFET VIFAVGREPQ LSKV LCETV GVKLDKNGRV VCTDDEQTTV SNVYAIGDIN AGKPQLTPVA IQAGRYLARR LFAGATELTD YSNVATTVFT PLEYG ACGL SEEDAIEKYG DKDIEVYHSN FKPLEWTVAH REDNVCYMKL VCRKSDNMRV LGLHVLGPNA GEITQGYAVA IKMGAT KAD FDRTIGIHPT CSETFTTLHV TKKSGVSPIV SGCCG

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Macromolecule #2: FLAVIN-ADENINE DINUCLEOTIDE

MacromoleculeName: FLAVIN-ADENINE DINUCLEOTIDE / type: ligand / ID: 2 / Number of copies: 2 / Formula: FAD
Molecular weightTheoretical: 785.55 Da
Chemical component information

ChemComp-FAD:
FLAVIN-ADENINE DINUCLEOTIDE / FAD*YM / Flavin adenine dinucleotide

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Macromolecule #3: (2~{R},3~{R},4~{S},5~{R})-2-[3-[[[(1~{R},2~{R},3~{R},5~{S})-2,6,6...

MacromoleculeName: (2~{R},3~{R},4~{S},5~{R})-2-[3-[[[(1~{R},2~{R},3~{R},5~{S})-2,6,6-trimethyl-3-bicyclo[3.1.1]heptanyl]amino]methyl]indol-1-yl]oxane-3,4,5-triol
type: ligand / ID: 3 / Number of copies: 2 / Formula: KW2
Molecular weightTheoretical: 414.538 Da
Chemical component information

ChemComp-KW2:
(2~{R},3~{R},4~{S},5~{R})-2-[3-[[[(1~{R},2~{R},3~{R},5~{S})-2,6,6-trimethyl-3-bicyclo[3.1.1]heptanyl]amino]methyl]indol-1-yl]oxane-3,4,5-triol

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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

Concentration0.4 mg/mL
BufferpH: 7.4
Component:
ConcentrationFormulaName
20.0 mMTRIStris(hydroxymethyl)aminomethane
50.0 mMNaClSodium chloridesodium chloride

Details: filtered aqueous fresh solution
GridModel: Quantifoil R1.2/1.3 / Material: GOLD / Mesh: 300 / Support film - Material: GOLD / Support film - topology: HOLEY ARRAY / Support film - Film thickness: 50 / Pretreatment - Type: GLOW DISCHARGE / Pretreatment - Time: 45 sec. / Pretreatment - Atmosphere: OTHER
VitrificationCryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 293.15 K / Instrument: FEI VITROBOT MARK IV / Details: Blotting time= 7s, wait time= 10 s.
DetailsMono-dispersed TGR molecules in aqueous sample buffer containing DMSO and inhibitor

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Electron microscopy

MicroscopeTFS GLACIOS
Electron beamAcceleration voltage: 200 kV / Electron source: FIELD EMISSION GUN
Electron opticsC2 aperture diameter: 150.0 µm / Illumination mode: SPOT SCAN / Imaging mode: DIFFRACTION / Cs: 2.7 mm / Nominal defocus max: 2.6 µm / Nominal defocus min: 1.8 µm / Nominal magnification: 12000
Sample stageCooling holder cryogen: NITROGEN
DetailsManual preliminar screening
Image recordingFilm or detector model: GATAN K2 SUMMIT (4k x 4k) / Digitization - Frames/image: 1-50 / Number grids imaged: 1 / Number real images: 2600 / Average electron dose: 50.0 e/Å2

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Image processing

Particle selectionNumber selected: 1479588
Startup modelType of model: NONE
Initial angle assignmentType: ANGULAR RECONSTITUTION / Software - Name: RELION
Final 3D classificationSoftware - Name: RELION
Final angle assignmentType: ANGULAR RECONSTITUTION / Software - Name: RELION
Final reconstructionApplied symmetry - Point group: C2 (2 fold cyclic) / Resolution.type: BY AUTHOR / Resolution: 3.6 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: RELION / Number images used: 173572
FSC plot (resolution estimation)

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Atomic model buiding 1

Initial modelPDB ID:

2v6o

RefinementSpace: REAL / Protocol: RIGID BODY FIT
Output model

PDB-8a1r:
cryo-EM structure of thioredoxin glutathione reductase in complex with a non-competitive inhibitor

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