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- PDB-8pty: Cryo-EM structure of human Elp123 in complex with 5'-deoxyadenosi... -

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Basic information

Entry
Database: PDB / ID: 8pty
TitleCryo-EM structure of human Elp123 in complex with 5'-deoxyadenosine and methionine
Components(Elongator complex protein ...) x 3
KeywordsTRANSLATION / Elongator / tRNA modification / acetyl-CoA hydrolysis
Function / homology
Function and homology information


phosphorylase kinase regulator activity / tRNA uridine(34) acetyltransferase activity / tRNA carboxymethyluridine synthase / elongator holoenzyme complex / tRNA wobble base 5-methoxycarbonylmethyl-2-thiouridinylation / tRNA wobble uridine modification / regulation of receptor signaling pathway via JAK-STAT / acetyltransferase activity / central nervous system development / transcription elongation factor complex ...phosphorylase kinase regulator activity / tRNA uridine(34) acetyltransferase activity / tRNA carboxymethyluridine synthase / elongator holoenzyme complex / tRNA wobble base 5-methoxycarbonylmethyl-2-thiouridinylation / tRNA wobble uridine modification / regulation of receptor signaling pathway via JAK-STAT / acetyltransferase activity / central nervous system development / transcription elongation factor complex / transcription elongation by RNA polymerase II / neuron migration / regulation of translation / HATs acetylate histones / 4 iron, 4 sulfur cluster binding / tRNA binding / positive regulation of cell migration / regulation of transcription by RNA polymerase II / protein kinase binding / nucleolus / nucleoplasm / metal ion binding / nucleus / cytoplasm / cytosol
Similarity search - Function
Elongator complex protein 1 / Elongator complex protein 2 / : / : / : / : / : / ELP1 first N-terminal beta-propeller / ELP1 N-terminal second beta-propeller / ELP1 TPR domain ...Elongator complex protein 1 / Elongator complex protein 2 / : / : / : / : / : / ELP1 first N-terminal beta-propeller / ELP1 N-terminal second beta-propeller / ELP1 TPR domain / ELP1 alpha-solenoid / ELP1 three-helix bundle / : / ELP3 N-terminal domain / Radical SAM, C-terminal extension / Elongator complex protein 3-like / ELP3/YhcC / Radical_SAM C-terminal domain / Elp3/MiaB/NifB / Elongator protein 3, MiaB family, Radical SAM / Radical SAM superfamily / Radical SAM core domain profile. / Radical SAM / Gcn5-related N-acetyltransferase (GNAT) domain profile. / GNAT domain / Acyl-CoA N-acyltransferase / WD domain, G-beta repeat / Trp-Asp (WD) repeats profile. / Trp-Asp (WD) repeats circular profile. / WD40 repeats / WD40 repeat / WD40-repeat-containing domain superfamily / WD40/YVTN repeat-like-containing domain superfamily
Similarity search - Domain/homology
5'-DEOXYADENOSINE / METHIONINE / IRON/SULFUR CLUSTER / Elongator complex protein 1 / Elongator complex protein 2 / Elongator complex protein 3
Similarity search - Component
Biological speciesHomo sapiens (human)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 3.58 Å
AuthorsAbbassi, N. / Jaciuk, M. / Lin, T.-Y. / Glatt, S.
Funding supportEuropean Union, 1items
OrganizationGrant numberCountry
European Research Council (ERC)101001394European Union
CitationJournal: Nat Commun / Year: 2024
Title: Cryo-EM structures of the human Elongator complex at work.
Authors: Nour-El-Hana Abbassi / Marcin Jaciuk / David Scherf / Pauline Böhnert / Alexander Rau / Alexander Hammermeister / Michał Rawski / Paulina Indyka / Grzegorz Wazny / Andrzej Chramiec- ...Authors: Nour-El-Hana Abbassi / Marcin Jaciuk / David Scherf / Pauline Böhnert / Alexander Rau / Alexander Hammermeister / Michał Rawski / Paulina Indyka / Grzegorz Wazny / Andrzej Chramiec-Głąbik / Dominika Dobosz / Bozena Skupien-Rabian / Urszula Jankowska / Juri Rappsilber / Raffael Schaffrath / Ting-Yu Lin / Sebastian Glatt /
Abstract: tRNA modifications affect ribosomal elongation speed and co-translational folding dynamics. The Elongator complex is responsible for introducing 5-carboxymethyl at wobble uridine bases (cmU) in ...tRNA modifications affect ribosomal elongation speed and co-translational folding dynamics. The Elongator complex is responsible for introducing 5-carboxymethyl at wobble uridine bases (cmU) in eukaryotic tRNAs. However, the structure and function of human Elongator remain poorly understood. In this study, we present a series of cryo-EM structures of human ELP123 in complex with tRNA and cofactors at four different stages of the reaction. The structures at resolutions of up to 2.9 Å together with complementary functional analyses reveal the molecular mechanism of the modification reaction. Our results show that tRNA binding exposes a universally conserved uridine at position 33 (U), which triggers acetyl-CoA hydrolysis. We identify a series of conserved residues that are crucial for the radical-based acetylation of U and profile the molecular effects of patient-derived mutations. Together, we provide the high-resolution view of human Elongator and reveal its detailed mechanism of action.
History
DepositionJul 16, 2023Deposition site: PDBE / Processing site: PDBE
Revision 1.0Apr 17, 2024Provider: repository / Type: Initial release
Revision 1.1May 22, 2024Group: Database references / Category: citation / citation_author
Item: _citation.country / _citation.journal_abbrev ..._citation.country / _citation.journal_abbrev / _citation.journal_id_CSD / _citation.journal_id_ISSN / _citation.journal_volume / _citation.page_first / _citation.page_last / _citation.pdbx_database_id_DOI / _citation.pdbx_database_id_PubMed / _citation.title / _citation.year

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

Downloads & links

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Assembly

Deposited unit
A: Elongator complex protein 1
B: Elongator complex protein 2
C: Elongator complex protein 3
hetero molecules


Theoretical massNumber of molelcules
Total (without water)309,5186
Polymers308,7663
Non-polymers7523
Water00
1


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

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Components

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Elongator complex protein ... , 3 types, 3 molecules ABC

#1: Protein Elongator complex protein 1 / ELP1 / IkappaB kinase complex-associated protein / IKK complex-associated protein / p150


Mass: 150427.484 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: ELP1, IKAP, IKBKAP / Production host: Spodoptera frugiperda (fall armyworm) / References: UniProt: O95163
#2: Protein Elongator complex protein 2 / ELP2 / SHINC-2 / STAT3-interacting protein 1 / StIP1


Mass: 92597.766 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: ELP2, STATIP1 / Production host: Spodoptera frugiperda (fall armyworm) / References: UniProt: Q6IA86
#3: Protein Elongator complex protein 3 / hELP3 / tRNA uridine(34) acetyltransferase


Mass: 65740.539 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: ELP3 / Production host: Spodoptera frugiperda (fall armyworm)
References: UniProt: Q9H9T3, Transferases; Acyltransferases; Transferring groups other than aminoacyl groups

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Non-polymers , 3 types, 3 molecules

#4: Chemical ChemComp-SF4 / IRON/SULFUR CLUSTER


Mass: 351.640 Da / Num. of mol.: 1 / Source method: obtained synthetically / Formula: Fe4S4 / Feature type: SUBJECT OF INVESTIGATION
#5: Chemical ChemComp-5AD / 5'-DEOXYADENOSINE


Mass: 251.242 Da / Num. of mol.: 1 / Source method: obtained synthetically / Formula: C10H13N5O3 / Feature type: SUBJECT OF INVESTIGATION
#6: Chemical ChemComp-MET / METHIONINE


Type: L-peptide linking / Mass: 149.211 Da / Num. of mol.: 1 / Source method: obtained synthetically / Formula: C5H11NO2S / Feature type: SUBJECT OF INVESTIGATION

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Details

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: Human Elp123 in complex with 5'-deoxyadenosine and methionine
Type: COMPLEX / Entity ID: #1-#3 / Source: RECOMBINANT
Molecular weightValue: 0.61 MDa / Experimental value: NO
Source (natural)Organism: Homo sapiens (human)
Source (recombinant)Organism: Spodoptera frugiperda (fall armyworm)
Buffer solutionpH: 7.5
Buffer component
IDConc.NameFormulaBuffer-ID
1100 mMsodium chlorideNaCl1
220 mMHEPESHEPES1
33 mMDithiothreitolDithiothreitol1
SpecimenConc.: 0.6 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
Specimen supportDetails: 8 mA / Grid material: COPPER / Grid mesh size: 200 divisions/in. / Grid type: Quantifoil R2/1
VitrificationInstrument: FEI VITROBOT MARK IV / Cryogen name: ETHANE / Humidity: 100 % / Chamber temperature: 277 K / Details: 15 s wait time, blot force 5, 5 s blot time

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Electron microscopy imaging

Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company
MicroscopyModel: TFS KRIOS
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Illumination mode: FLOOD BEAM
Electron lensMode: BRIGHT FIELD / Nominal magnification: 96000 X / Nominal defocus max: 2700 nm / Nominal defocus min: 900 nm / Cs: 2.7 mm / C2 aperture diameter: 50 µm / Alignment procedure: COMA FREE
Specimen holderCryogen: NITROGEN / Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER
Image recordingElectron dose: 40 e/Å2 / Detector mode: COUNTING / Film or detector model: FEI FALCON III (4k x 4k) / Num. of real images: 5300
Image scansWidth: 4096 / Height: 4096

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Processing

EM software
IDNameCategory
1cryoSPARCparticle selection
2Topazparticle selection
3EPUimage acquisition
5cryoSPARCCTF correction
8UCSF ChimeraXmodel fitting
9NAMDmodel fitting
11cryoSPARCinitial Euler assignment
12RELIONfinal Euler assignment
13RELIONclassification
14RELION3D reconstruction
15Cootmodel refinement
16PHENIXmodel refinement
CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
Particle selectionNum. of particles selected: 264351
Details: given number of particles from TOPAZ picking, and after 3D curation
3D reconstructionResolution: 3.58 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 171951 / Symmetry type: POINT
Atomic model buildingDetails: Based on PDB 6QK7 / Source name: SwissModel / Type: in silico model
Refine LS restraints
Refine-IDTypeDev idealNumber
ELECTRON MICROSCOPYf_bond_d0.00414551
ELECTRON MICROSCOPYf_angle_d0.59719777
ELECTRON MICROSCOPYf_dihedral_angle_d4.681917
ELECTRON MICROSCOPYf_chiral_restr0.0452201
ELECTRON MICROSCOPYf_plane_restr0.0042527

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