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- PDB-8qzx: CryoEM structure of DHS-eIF5A complex structure from Trichomonas ... -

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Basic information

Entry
Database: PDB / ID: 8qzx
TitleCryoEM structure of DHS-eIF5A complex structure from Trichomonas vaginalis
Components
  • Deoxyhypusine synthase related protein, putative
  • Eukaryotic translation initiation factor 5A
KeywordsTRANSLATION / parasite / hypusination / eI5FA / hypusine / DHS / Trichomonas vaginalis / deoxyhypusination / translation factor
Function / homology
Function and homology information


deoxyhypusine synthase activity / positive regulation of translational termination / positive regulation of translational elongation / translation elongation factor activity / translation initiation factor activity / ribosome binding / RNA binding / cytoplasm
Similarity search - Function
Deoxyhypusine synthase / Deoxyhypusine synthase superfamily / Deoxyhypusine synthase / : / Translation initiation factor 5A-like, N-terminal / Translation elongation factor, IF5A, hypusine site / Eukaryotic initiation factor 5A hypusine signature. / Eukaryotic elongation factor 5A hypusine, DNA-binding OB fold / Translation elongation factor IF5A-like / Translation elongation factor, IF5A C-terminal ...Deoxyhypusine synthase / Deoxyhypusine synthase superfamily / Deoxyhypusine synthase / : / Translation initiation factor 5A-like, N-terminal / Translation elongation factor, IF5A, hypusine site / Eukaryotic initiation factor 5A hypusine signature. / Eukaryotic elongation factor 5A hypusine, DNA-binding OB fold / Translation elongation factor IF5A-like / Translation elongation factor, IF5A C-terminal / Eukaryotic elongation factor 5A hypusine, DNA-binding OB fold / DHS-like NAD/FAD-binding domain superfamily / Translation protein SH3-like domain superfamily / Ribosomal protein L2, domain 2 / Nucleic acid-binding, OB-fold
Similarity search - Domain/homology
NICOTINAMIDE-ADENINE-DINUCLEOTIDE / SPERMIDINE / Deoxyhypusine synthase related protein, putative / Eukaryotic translation initiation factor 5A
Similarity search - Component
Biological speciesTrichomonas vaginalis (eukaryote)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 3.01 Å
AuthorsWator, E. / Wilk, P. / Grudnik, P.
Funding support Poland, 2items
OrganizationGrant numberCountry
Polish National Science Centre2019/35/N/NZ1/02805 Poland
Polish National Science Centre2019/33/B/NZ1/01839 Poland
CitationJournal: FEBS J / Year: 2024
Title: Structural characterization of the (deoxy)hypusination in Trichomonas vaginalis questions the bifunctionality of deoxyhypusine synthase.
Authors: Elżbieta Wątor / Piotr Wilk / Paweł Kochanowski / Przemysław Grudnik /
Abstract: Trichomonas vaginalis, the causative agent of trichomoniasis, is a prevalent anaerobic protozoan parasite responsible for the most common nonviral sexually transmitted infection globally. While ...Trichomonas vaginalis, the causative agent of trichomoniasis, is a prevalent anaerobic protozoan parasite responsible for the most common nonviral sexually transmitted infection globally. While metronidazole and its derivatives are approved drugs for this infection, rising resistance necessitates the exploration of new antiparasitic therapies. Protein posttranslational modifications (PTMs) play crucial roles in cellular processes, and among them, hypusination, involving eukaryotic translation factor 5A (eIF5A), has profound implications. Despite extensive studies in various organisms, the role of hypusination in T. vaginalis and its potential impact on parasite biology and pathogenicity remain poorly understood. This study aims to unravel the structural basis of the hypusination pathway in T. vaginalis using X-ray crystallography and cryo-electron microscopy. The results reveal high structural homology between T. vaginalis and human orthologs, providing insights into the molecular architecture of eIF5A and deoxyhypusine synthase (DHS) and their interaction. Contrary to previous suggestions of bifunctionality, our analyses indicate that the putative hydroxylation site in tvDHS is nonfunctional, and biochemical assays demonstrate exclusive deoxyhypusination capability. These findings challenge the notion of tvDHS functioning as both deoxyhypusine synthase and hydroxylase. The study enhances understanding of the hypusination pathway in T. vaginalis, shedding light on its functional relevance and potential as a drug target, and contributing to the development of novel therapeutic strategies against trichomoniasis.
History
DepositionOct 30, 2023Deposition site: PDBE / Processing site: PDBE
Revision 1.0Jul 3, 2024Provider: repository / Type: Initial release
Revision 1.1Jul 10, 2024Group: Data collection / Database references / Category: citation / citation_author / em_admin
Item: _citation.pdbx_database_id_PubMed / _citation.title ..._citation.pdbx_database_id_PubMed / _citation.title / _citation_author.identifier_ORCID / _em_admin.last_update

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

Downloads & links

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Assembly

Deposited unit
A: Deoxyhypusine synthase related protein, putative
B: Deoxyhypusine synthase related protein, putative
C: Deoxyhypusine synthase related protein, putative
D: Deoxyhypusine synthase related protein, putative
E: Eukaryotic translation initiation factor 5A
hetero molecules


Theoretical massNumber of molelcules
Total (without water)183,47612
Polymers180,3875
Non-polymers3,0897
Water00
1


  • Idetical with deposited unit
  • defined by author
  • Evidence: electron microscopy, not applicable, gel filtration
TypeNameSymmetry operationNumber
identity operation1_5551

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Components

#1: Protein
Deoxyhypusine synthase related protein, putative


Mass: 40489.102 Da / Num. of mol.: 4 / Mutation: K332A
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Trichomonas vaginalis (eukaryote) / Gene: TVAG_359990 / Production host: Escherichia coli BL21(DE3) (bacteria) / Variant (production host): Rosetta / References: UniProt: A2DTB8
#2: Protein Eukaryotic translation initiation factor 5A / eIF-5A


Mass: 18430.594 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Trichomonas vaginalis (eukaryote) / Gene: eIF-5A1 / Production host: Escherichia coli (E. coli) / References: UniProt: D5MC19
#3: Chemical
ChemComp-NAD / NICOTINAMIDE-ADENINE-DINUCLEOTIDE


Mass: 663.425 Da / Num. of mol.: 4 / Source method: obtained synthetically / Formula: C21H27N7O14P2 / Feature type: SUBJECT OF INVESTIGATION / Comment: NAD*YM
#4: Chemical ChemComp-SPD / SPERMIDINE / N-(2-AMINO-PROPYL)-1,4-DIAMINOBUTANE / PA(34)


Mass: 145.246 Da / Num. of mol.: 3 / Source method: obtained synthetically / Formula: C7H19N3 / Feature type: SUBJECT OF INVESTIGATION
Has ligand of interestY

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Experimental details

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Experiment

ExperimentMethod: ELECTRON MICROSCOPY
EM experimentAggregation state: PARTICLE / 3D reconstruction method: single particle reconstruction

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Sample preparation

ComponentName: Complex of DHS-eIF5A / Type: COMPLEX / Entity ID: #1-#2 / Source: RECOMBINANT
Molecular weightExperimental value: NO
Source (natural)Organism: Trichomonas vaginalis (eukaryote)
Source (recombinant)Organism: Escherichia coli (E. coli)
Buffer solutionpH: 9.3
Buffer component
IDConc.NameFormulaBuffer-ID
1200 mMglycine bufferNaOH/Glycine1
2200 mMsodium chlorideNaCl1
SpecimenConc.: 0.5 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
Specimen supportGrid type: Quantifoil R2/2
VitrificationInstrument: FEI VITROBOT MARK IV / Cryogen name: ETHANE / Humidity: 100 % / Chamber temperature: 278 K

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Electron microscopy imaging

Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company
MicroscopyModel: FEI TITAN KRIOS
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Illumination mode: FLOOD BEAM
Electron lensMode: BRIGHT FIELD / Nominal defocus max: 3500 nm / Nominal defocus min: 500 nm / Cs: 2.7 mm
Specimen holderCryogen: NITROGEN
Image recordingElectron dose: 40 e/Å2 / Film or detector model: GATAN K3 (6k x 4k)

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Processing

EM software
IDNameVersionCategory
1cryoSPARCparticle selection
2EPUimage acquisition
7UCSF ChimeraXmodel fitting
12cryoSPARC3D reconstruction
13PHENIX1.20.1_4487:model refinement
Image processingDetails: selected images were normalized and low-pass filtered
CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
Particle selectionNum. of particles selected: 1157408 / Details: template picking
SymmetryPoint symmetry: C1 (asymmetric)
3D reconstructionResolution: 3.01 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 422161 / Algorithm: FOURIER SPACE / Num. of class averages: 1 / Symmetry type: POINT
Atomic model buildingProtocol: RIGID BODY FIT / Space: REAL
Atomic model buildingPDB-ID: 8A0E

8a0e


Accession code: 8A0E / Source name: PDB / Type: experimental model
Refine LS restraints
Refine-IDTypeDev idealNumber
ELECTRON MICROSCOPYf_bond_d0.00312285
ELECTRON MICROSCOPYf_angle_d0.86416687
ELECTRON MICROSCOPYf_dihedral_angle_d11.531760
ELECTRON MICROSCOPYf_chiral_restr0.0491844
ELECTRON MICROSCOPYf_plane_restr0.0062140

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