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- EMDB-18397: 5'vRNA-bound Hantaan virus polymerase in monomeric active state -

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Basic information

Entry
Database: EMDB / ID: EMD-18397
Title5'vRNA-bound Hantaan virus polymerase in monomeric active state
Map data
Sample
  • Complex: Hantaan virus polymerase
    • Complex: RNA-directed RNA polymerase L
      • Protein or peptide: RNA-directed RNA polymerase L
    • Complex: RNA (5'-R(P*AP*GP*UP*AP*GP*UP*AP*GP*AP*CP*A)-3')
      • RNA: RNA (5'-R(P*AP*GP*UP*AP*GP*UP*AP*GP*AP*CP*A)-3')
  • Ligand: MAGNESIUM ION
KeywordsBunyavirus / Hantaan virus / polymerase / dimer / VIRAL PROTEIN
Function / homology
Function and homology information


RNA-templated viral transcription / negative stranded viral RNA replication / cap snatching / endonuclease activity / Hydrolases; Acting on ester bonds / host cell perinuclear region of cytoplasm / RNA-directed RNA polymerase / RNA-dependent RNA polymerase activity / nucleotide binding / DNA-templated transcription / metal ion binding
Similarity search - Function
RNA-directed RNA polymerase, hantavirus / RNA-directed RNA polymerase, hantavirus, N-terminal / RNA dependent RNA polymerase / : / RNA-dependent RNA polymerase, bunyaviral / Bunyavirus RNA dependent RNA polymerase / RNA-directed RNA polymerase, negative-strand RNA virus / RdRp of negative ssRNA viruses with segmented genomes catalytic domain profile.
Similarity search - Domain/homology
RNA-directed RNA polymerase L
Similarity search - Component
Biological speciesHantaan virus 76-118
Methodsingle particle reconstruction / cryo EM / Resolution: 2.81 Å
AuthorsDurieux Trouilleton Q / Arragain B / Malet H
Funding support France, 1 items
OrganizationGrant numberCountry
Agence Nationale de la Recherche (ANR)ANR-19-CE11-0024 France
CitationJournal: Nat Commun / Year: 2024
Title: Structural characterization of the oligomerization of full-length Hantaan virus polymerase into symmetric dimers and hexamers.
Authors: Quentin Durieux Trouilleton / Dominique Housset / Paco Tarillon / Benoît Arragain / Hélène Malet /
Abstract: Hantaan virus is a dangerous human pathogen whose segmented negative-stranded RNA genome is replicated and transcribed by a virally-encoded multi-functional polymerase. Here we describe the complete ...Hantaan virus is a dangerous human pathogen whose segmented negative-stranded RNA genome is replicated and transcribed by a virally-encoded multi-functional polymerase. Here we describe the complete cryo-electron microscopy structure of Hantaan virus polymerase in several oligomeric forms. Apo polymerase protomers can adopt two drastically different conformations, which assemble into two distinct symmetric homodimers, that can themselves gather to form hexamers. Polymerase dimerization induces the stabilization of most polymerase domains, including the C-terminal domain that contributes the most to dimer's interface, along with a lariat region that participates to the polymerase steadying. Binding to viral RNA induces significant conformational changes resulting in symmetric oligomer disruption and polymerase activation, suggesting the possible involvement of apo multimers as protecting systems that would stabilize the otherwise flexible C-terminal domains. Overall, these results provide insights into the multimerization capability of Hantavirus polymerase and may help to define antiviral compounds to counteract these life-threatening viruses.
History
DepositionSep 6, 2023-
Header (metadata) releaseMar 27, 2024-
Map releaseMar 27, 2024-
UpdateMar 27, 2024-
Current statusMar 27, 2024Processing site: PDBe / Status: Released

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Structure visualization

Supplemental images

emd_18397.png

Downloads & links

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Map

FileDownload / File: emd_18397.map.gz / Format: CCP4 / Size: 166.4 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Voxel sizeX=Y=Z: 0.839 Å
Density
Contour LevelBy AUTHOR: 0.385
Minimum - Maximum-3.1992114 - 4.7787395
Average (Standard dev.)0.0014568528 (±0.06488377)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions352352352
Spacing352352352
CellA=B=C: 295.328 Å
α=β=γ: 90.0 °

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Supplemental data

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Half map: #2

Fileemd_18397_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_18397_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 : Hantaan virus polymerase

EntireName: Hantaan virus polymerase
Components
  • Complex: Hantaan virus polymerase
    • Complex: RNA-directed RNA polymerase L
      • Protein or peptide: RNA-directed RNA polymerase L
    • Complex: RNA (5'-R(P*AP*GP*UP*AP*GP*UP*AP*GP*AP*CP*A)-3')
      • RNA: RNA (5'-R(P*AP*GP*UP*AP*GP*UP*AP*GP*AP*CP*A)-3')
  • Ligand: MAGNESIUM ION

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Supramolecule #1: Hantaan virus polymerase

SupramoleculeName: Hantaan virus polymerase / type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1-#2
Molecular weightTheoretical: 250 KDa

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Supramolecule #2: RNA-directed RNA polymerase L

SupramoleculeName: RNA-directed RNA polymerase L / type: complex / ID: 2 / Parent: 1 / Macromolecule list: #1
Source (natural)Organism: Hantaan virus 76-118

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Supramolecule #3: RNA (5'-R(P*AP*GP*UP*AP*GP*UP*AP*GP*AP*CP*A)-3')

SupramoleculeName: RNA (5'-R(P*AP*GP*UP*AP*GP*UP*AP*GP*AP*CP*A)-3') / type: complex / ID: 3 / Parent: 1 / Macromolecule list: #2
Source (natural)Organism: Hantaan virus 76-118

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Macromolecule #1: RNA-directed RNA polymerase L

MacromoleculeName: RNA-directed RNA polymerase L / type: protein_or_peptide / ID: 1 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Hantaan virus 76-118
Molecular weightTheoretical: 249.476484 KDa
Recombinant expressionOrganism: Trichoplusia ni (cabbage looper)
SequenceString: MGHHHHHHDY DIPTTENLYF QGMDKYREIH NKLKEFSPGT LTAVECIDYL DRLYAVRHDI VDQMIKHDWS DNKDSEEAIG KVLLFAGVP SNIITALEKK IIPNHPTGKS LKAFFKMTPD NYKISGTTIE FVEVTVTADV DKGIREKKLK YEAGLTYIEQ E LHKFFLKG ...String:
MGHHHHHHDY DIPTTENLYF QGMDKYREIH NKLKEFSPGT LTAVECIDYL DRLYAVRHDI VDQMIKHDWS DNKDSEEAIG KVLLFAGVP SNIITALEKK IIPNHPTGKS LKAFFKMTPD NYKISGTTIE FVEVTVTADV DKGIREKKLK YEAGLTYIEQ E LHKFFLKG EIPQPYKITF NVVAVRTDGS NITTQWPSRR NDGVVQYMRL VQAEISYVRE HLIKTEERAA LEAMFNLKFN IS THKSQPY YIPDYKGMEP IGANIEDLVD YSKDWLSRAR NFSFFEVKGT AVFECFNSNE ANHCQRYPMS RKPRNFLLIQ CSL ITSYKP ATTLSDQIDS RRACSYILNL IPDTPASYLI HDMAYRYINL TREDMINYYA PRIQFKQTQN VREPGTFKLT SSML RAESK AMLDLLNNHK SGEKHGAQIE SLNIASHIVQ SESVSLITKI LSDLELNITE PSTQEYSTTK HTYVDTVLDK FFQNE TQKY LIDVLKKTTA WHIGHLIRDI TESLIAHSGL KRSKYWSLHS YNNGNVILFI LPSKSLEVAG SFIRFITVFR IGPGLV DKD NLDTILIDGD SQWGVSKVMS IDLNRLLALN IAFEKALIAT ATWFQYYTED QGQFPLQYAI RSVFANHFLL AICQKMK LC AIFDNLRYLI PAVTSLYSGF PSLIEKLFER PFKSSLEVYI YYNIKSLLVA LAQNNKARFY SKVKLLGLTV DQSTVGAS G VYPSFMSRIV YKHYRSLISE VTTCFFLFEK GLHGNMNEEA KIHLETVEWA LKFREKEEKY GESLVENGYM MWELRANAE LAEQQLYCQD AIELAAIELN KVLATKSSVV ANSILSKNWE EPYFSQTRNI SLKGMSGQVQ EDGHLSSSVT IIEAIRYLSN SRHNPSLLK LYEETREQKA MARIVRKYQR TEADRGFFIT TLPTRCRLEI IEDYYDAIAK NISEEYISYG GEKKILAIQG A LEKALRWA SGESFIELSN HKFIRMKRKL MYVSADATKW SPGDNSAKFR RFTSMLHNGL PNNKLKNCVI DALKQVYKTD FF MSRKLRN YIDSMESLDP HIKQFLDFFP DGHHGEVKGN WLQGNLNKCS SLFGVAMSLL FKQVWTNLFP ELDCFFEFAH HSD DALFIY GYLEPVDDGT DWFLFVSQQI QAGHLHWFSV NTEMWKSMFN LHEHILLLGS IKISPKKTTV SPTNAEFLST FFEG CAVSI PFVKILLGSL SDLPGLGYFD DLAAAQSRCV KALDLGASPQ VAQLAVALCT SKVERLYGTA PGMVNHPAAY LQVKH TDTP IPLGGNGAMS IMELATAGIG MSDKNLLKRA LLGYSHKRQK SMLYILGLFK FLMKLSDETF QHERLGQFSF IGKVQW KIF TPKSEFEFAD MYTSKFLELW SSQHVTYDYI IPKGRDNLLI YLVRKLNDPS IVTAMTMQSP LQLRFRMQAK QHMKVCR LD GEWVTFREVL AAANSFAENY SATSQDMDLF QTLTSCTFSK EYAWKDFLNG IHCDVIPTKQ VQRAKVARTF TVREKDQI I QNSIPAVIGY KFAVTVEEMS DVLDTAKFPD SLSVDLKTMK DGVYRELGLD ISLPDVMKRI APMLYKSSKS RVVIVQGNV EGTAEAICRY WLKSMSLVKT IRVKPHKEVL QAVSIFNRKE DIGQQKDLAA LKLCIEVWRW CKANSAPYRD WFQALWFEDK TFSEWLDRF CRVGVPPIDP EIQCAALMIA DIKGDYSVLQ LQANRRAYSG KQYDAYCVQT YNEVTKLYEG DLRVTFNFGL D CARLEIFW DKKAYILETS ITQKHVLKIM MDEVSKELIK CGMRFNTEQV QGVRHMVLFK TESGFEWGKP NIPCIVYKNC VL RTSLRTT QAINHKFMIT IKDDGLRAIA QHDEDSPRFL LAHAFHTIRD IRYQAVDAVS NVWFIHKGVK LYLNPIISSG LLE NFMKNL PAAIPPAAYS LIMNRAKISV DLFMFNDLLK LINPRNTLDL SGLETTGDEF STVSSMSSRL WSEEMSLVDD DEEL DDEFT IDLQDVDFEN IDIEADIEHF LQDESSYTGD LLISTEETES KKMRGIVKIL EPVRLIKSWV SRGLSIEKVY SPVNI ILMS RYISKTFNLS TKQVSLLDPY DLTELESIVR GWGECVIDQF ESLDREAQNM VVNKGICPED VIPDSLFSFR HTMVLL RRL FPQDSISSFY

UniProtKB: RNA-directed RNA polymerase L

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Macromolecule #2: RNA (5'-R(P*AP*GP*UP*AP*GP*UP*AP*GP*AP*CP*A)-3')

MacromoleculeName: RNA (5'-R(P*AP*GP*UP*AP*GP*UP*AP*GP*AP*CP*A)-3') / type: rna / ID: 2 / Number of copies: 1
Source (natural)Organism: Hantaan virus 76-118
Molecular weightTheoretical: 8.046852 KDa
SequenceString:
UAGUAGUAGA CACCGCAAGA UGUUA

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Macromolecule #3: MAGNESIUM ION

MacromoleculeName: MAGNESIUM ION / type: ligand / ID: 3 / Number of copies: 1 / Formula: MG
Molecular weightTheoretical: 24.305 Da

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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

Concentration0.25 mg/mL
BufferpH: 8
Component:
ConcentrationFormulaName
30.0 mMC8H18N2O4SHEPES
250.0 mMNaClSodium chlorideNaClSodium chloride
10.0 mMC9H15O6PTCEP
GridModel: UltrAuFoil R1.2/1.3 / Material: GOLD / Mesh: 300 / Support film - Material: GOLD / Support film - topology: HOLEY / Pretreatment - Type: GLOW DISCHARGE / Pretreatment - Time: 50 sec. / Pretreatment - Atmosphere: AIR
VitrificationCryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 77 K / Instrument: FEI VITROBOT MARK IV / Details: blot force 1 3s.

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsCalibrated defocus max: 2.0 µm / Calibrated defocus min: 0.8 µm / Calibrated magnification: 105000 / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELDBright-field microscopy / Nominal defocus max: 2.0 µm / Nominal defocus min: 0.8 µm / Nominal magnification: 105000
Specialist opticsEnergy filter - Name: GIF Quantum LS / Energy filter - Slit width: 20 eV
Sample stageSpecimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Cooling holder cryogen: NITROGEN
Image recordingFilm or detector model: GATAN K3 (6k x 4k) / Digitization - Dimensions - Width: 5760 pixel / Digitization - Dimensions - Height: 4092 pixel / Number grids imaged: 1 / Number real images: 26745 / Average exposure time: 1.45 sec. / Average electron dose: 40.0 e/Å2
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Particle selectionNumber selected: 3285016
Startup modelType of model: INSILICO MODEL / In silico model: Ab-initio reconstruction cryoSPARC
Initial angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: cryoSPARC (ver. 4.2.1)
Final 3D classificationNumber classes: 5 / Software - Name: cryoSPARC (ver. 4.2.1)
Final angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: cryoSPARC (ver. 4.2.1)
Final reconstructionResolution.type: BY AUTHOR / Resolution: 2.81 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: cryoSPARC (ver. 4.2.1) / Number images used: 882132
FSC plot (resolution estimation)

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Atomic model buiding 1

Initial modelPDB ID:

8c4s


Chain - Chain ID: A / Chain - Source name: PDB / Chain - Initial model type: experimental model
RefinementSpace: REAL / Protocol: AB INITIO MODEL / Overall B value: 8.45
Output model

PDB-8qh3:
5'vRNA-bound Hantaan virus polymerase in monomeric active state

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