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- EMDB-39020: FluPol-NS2 complex (local refinement) -

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Basic information

Entry
Database: EMDB / ID: EMD-39020
TitleFluPol-NS2 complex (local refinement)
Map data
Sample
  • Complex: FluPol-NS2 complex (local refinement)
    • Complex: FluPol
      • Protein or peptide: Polymerase acidic protein
      • Protein or peptide: RNA-directed RNA polymerase catalytic subunit
      • Protein or peptide: Polymerase basic protein 2
    • Complex: NS2
      • Protein or peptide: Nuclear export protein
KeywordsPROTEIN BINDING
Function / homology
Function and homology information


exit of virus from host cell nucleus through nuclear pore / cap snatching / viral transcription / symbiont-mediated suppression of host mRNA transcription via inhibition of RNA polymerase II activity / 7-methylguanosine mRNA capping / host cell mitochondrion / symbiont-mediated suppression of host cytoplasmic pattern recognition receptor signaling pathway via inhibition of MAVS activity / virion component / endonuclease activity / Hydrolases; Acting on ester bonds ...exit of virus from host cell nucleus through nuclear pore / cap snatching / viral transcription / symbiont-mediated suppression of host mRNA transcription via inhibition of RNA polymerase II activity / 7-methylguanosine mRNA capping / host cell mitochondrion / symbiont-mediated suppression of host cytoplasmic pattern recognition receptor signaling pathway via inhibition of MAVS activity / virion component / endonuclease activity / Hydrolases; Acting on ester bonds / host cell cytoplasm / symbiont-mediated suppression of host gene expression / viral translational frameshifting / RNA-directed RNA polymerase / viral RNA genome replication / RNA-directed RNA polymerase activity / nucleotide binding / DNA-templated transcription / host cell nucleus / RNA binding / metal ion binding
Similarity search - Function
Influenza nuclear export protein NS2 / Influenza non-structural protein (NS2) / Influenza RNA-dependent RNA polymerase subunit PB2 / PB2, C-terminal / Influenza RNA-dependent RNA polymerase subunit PB1 / Influenza RNA-dependent RNA polymerase subunit PB1 / : / : / Influenza RNA polymerase PB2 N-terminal region / Influenza RNA polymerase PB2 second domain ...Influenza nuclear export protein NS2 / Influenza non-structural protein (NS2) / Influenza RNA-dependent RNA polymerase subunit PB2 / PB2, C-terminal / Influenza RNA-dependent RNA polymerase subunit PB1 / Influenza RNA-dependent RNA polymerase subunit PB1 / : / : / Influenza RNA polymerase PB2 N-terminal region / Influenza RNA polymerase PB2 second domain / : / Influenza RNA polymerase PB2 middle domain / : / Influenza RNA polymerase PB2 C-terminal domain / : / Influenza RNA polymerase PB2 6th domain / : / Influenza RNA polymerase PB2 CAP binding domain / : / Influenza RNA polymerase PB2 helical domain / Polymerase acidic protein / Influenza RNA-dependent RNA polymerase subunit PA / Influenza RNA-dependent RNA polymerase subunit PA, endonuclease domain / Influenza RNA-dependent RNA polymerase subunit PA / RNA-directed RNA polymerase, negative-strand RNA virus / RdRp of negative ssRNA viruses with segmented genomes catalytic domain profile.
Similarity search - Domain/homology
Polymerase basic protein 2 / Polymerase acidic protein / RNA-directed RNA polymerase catalytic subunit / Nuclear export protein
Similarity search - Component
Biological speciesInfluenza A virus (A/Aichi/2/1968(H3N2)) / Influenza A virus (A/nt/60/1968(H3N2)) / Influenza A virus (strain A/Hong Kong/1/1968 H3N2)
Methodsingle particle reconstruction / cryo EM / Resolution: 3.0 Å
AuthorsPeng Q / Sun JQ
Funding support China, 2 items
OrganizationGrant numberCountry
National Natural Science Foundation of China (NSFC)81871658 China
National Natural Science Foundation of China (NSFC)32192452 China
CitationJournal: EMBO Rep / Year: 2024
Title: NS2 induces an influenza A RNA polymerase hexamer and acts as a transcription to replication switch.
Authors: Junqing Sun / Lu Kuai / Lei Zhang / Yufeng Xie / Yanfang Zhang / Yan Li / Qi Peng / Yuekun Shao / Qiuxian Yang / Wen-Xia Tian / Junhao Zhu / Jianxun Qi / Yi Shi / Tao Deng / George F Gao /
Abstract: Genome transcription and replication of influenza A virus (FluA), catalyzed by viral RNA polymerase (FluAPol), are delicately controlled across the virus life cycle. A switch from transcription to ...Genome transcription and replication of influenza A virus (FluA), catalyzed by viral RNA polymerase (FluAPol), are delicately controlled across the virus life cycle. A switch from transcription to replication occurring at later stage of an infection is critical for progeny virion production and viral non-structural protein NS2 has been implicated in regulating the switch. However, the underlying regulatory mechanisms and the structure of NS2 remained elusive for years. Here, we determine the cryo-EM structure of the FluAPol-NS2 complex at ~3.0 Å resolution. Surprisingly, three domain-swapped NS2 dimers arrange three symmetrical FluPol dimers into a highly ordered barrel-like hexamer. Further structural and functional analyses demonstrate that NS2 binding not only hampers the interaction between FluAPol and the Pol II CTD because of steric conflicts, but also impairs FluAPol transcriptase activity by stalling it in the replicase conformation. Moreover, this is the first visualization of the full-length NS2 structure. Our findings uncover key molecular mechanisms of the FluA transcription-replication switch and have implications for the development of antivirals.
History
DepositionFeb 4, 2024-
Header (metadata) releaseJul 24, 2024-
Map releaseJul 24, 2024-
UpdateDec 11, 2024-
Current statusDec 11, 2024Processing site: PDBj / Status: Released

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Structure visualization

Supplemental images

emd_39020.png

Downloads & links

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Map

FileDownload / File: emd_39020.map.gz / Format: CCP4 / Size: 669.9 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
0.89 Å/pix.
x 560 pix.
= 498.4 Å		0.89 Å/pix.
x 560 pix.
= 498.4 Å		0.89 Å/pix.
x 560 pix.
= 498.4 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 0.89 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.012
Minimum - Maximum-0.0019749883 - 1.6164823
Average (Standard dev.)0.00055134104 (±0.016631842)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions560560560
Spacing560560560
CellA=B=C: 498.4 Å
α=β=γ: 90.0 °

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Supplemental data

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Half map: #2

Fileemd_39020_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_39020_half_map_2.map
Projections & Slices
AxesZYX

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Histogram (log scale)

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Sample components

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Entire : FluPol-NS2 complex (local refinement)

EntireName: FluPol-NS2 complex (local refinement)
Components
  • Complex: FluPol-NS2 complex (local refinement)
    • Complex: FluPol
      • Protein or peptide: Polymerase acidic protein
      • Protein or peptide: RNA-directed RNA polymerase catalytic subunit
      • Protein or peptide: Polymerase basic protein 2
    • Complex: NS2
      • Protein or peptide: Nuclear export protein

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Supramolecule #1: FluPol-NS2 complex (local refinement)

SupramoleculeName: FluPol-NS2 complex (local refinement) / type: complex / ID: 1 / Parent: 0 / Macromolecule list: all

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Supramolecule #2: FluPol

SupramoleculeName: FluPol / type: complex / ID: 2 / Parent: 1 / Macromolecule list: #1-#3
Source (natural)Organism: Influenza A virus (A/Aichi/2/1968(H3N2))

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Supramolecule #3: NS2

SupramoleculeName: NS2 / type: complex / ID: 3 / Parent: 1 / Macromolecule list: #4
Source (natural)Organism: Influenza A virus (A/Aichi/2/1968(H3N2))

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Macromolecule #1: Polymerase acidic protein

MacromoleculeName: Polymerase acidic protein / type: protein_or_peptide / ID: 1 / Number of copies: 2 / Enantiomer: LEVO / EC number: Hydrolases; Acting on ester bonds
Source (natural)Organism: Influenza A virus (A/nt/60/1968(H3N2)) / Strain: A/NT/60/68/(H3N2)
Molecular weightTheoretical: 84.625414 KDa
Recombinant expressionOrganism: Homo sapiens (human)
SequenceString: MHHHHHHHHG SGSMEDFVRQ CFNPMIVELA EKAMKEYGED LKIETNKFAA ICTHLEVCFM YSDFHFINEQ GESIVVELDD PNALLKHRF EIIEGRDRTM AWTVVNSICN TTGAEKPKFL PDLYDYKENR FIEIGVTRRE VHIYYLEKAN KIKSENTHIH I FSFTGEEM ...String:
MHHHHHHHHG SGSMEDFVRQ CFNPMIVELA EKAMKEYGED LKIETNKFAA ICTHLEVCFM YSDFHFINEQ GESIVVELDD PNALLKHRF EIIEGRDRTM AWTVVNSICN TTGAEKPKFL PDLYDYKENR FIEIGVTRRE VHIYYLEKAN KIKSENTHIH I FSFTGEEM ATKADYTLDE ESRARIKTRL FTIRQEMANR GLWDSFRQSE RGEETIEERF EITGTMRRLA DQSLPPNFSC LE NFRAYVD GFEPNGYIEG KLSQMSKEVN AKIEPFLKTT PRPIRLPDGP PCFQRSKFLL MDALKLSIED PSHEGEGIPL YDA IKCMRT FFGWKEPYIV KPHEKGINPN YLLSWKQVLA ELQDIENEEK IPRTKNMKKT SQLKWALGEN MAPEKVDFDN CRDV SDLKQ YDSDEPELRS LSSWIQNEFN KACELTDSTW IELDEIGEDV APIEYIASMR RNYFTAEVSH CRATEYIMKG VYINT ALLN ASCAAMDDFQ LIPMISKCRT KEGRRKTNLY GFIIKGRSHL RNDTDVVNFV SMEFSLTDPR LEPHKWEKYC VLEIGD MLL RSAIGQMSRP MFLYVRTNGT SKIKMKWGME MRRCLLQSLQ QIESMIEAES SVKEKDMTKE FFENKSETWP IGESPKG VE DGSIGKVCRT LLAKSVFNSL YASPQLEGFS AESRKLLLVV QALRDNLEPG TFDLEGLYEA IEECLINDPW VLLNASWF N SFLTHALR

UniProtKB: Polymerase acidic protein

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Macromolecule #2: RNA-directed RNA polymerase catalytic subunit

MacromoleculeName: RNA-directed RNA polymerase catalytic subunit / type: protein_or_peptide / ID: 2 / Number of copies: 2 / Enantiomer: LEVO / EC number: RNA-directed RNA polymerase
Source (natural)Organism: Influenza A virus (strain A/Hong Kong/1/1968 H3N2)
Strain: A/Hong Kong/1/1968 H3N2
Molecular weightTheoretical: 86.524086 KDa
Recombinant expressionOrganism: Homo sapiens (human)
SequenceString: MDVNPTLLFL KVPAQNAIST TFPYTGDPPY SHGTGTGYTM DTVNRTHQYS EKGKWTTNTE TGAPQLNPID GPLPEDNEPS GYAQTDCVL EAMAFLEESH PGIFENSCLE TMEVVQQTRV DRLTQGRQTY DWTLNRNQPA ATALANTIEV FRSNGLTANE S GRLIDFLK ...String:
MDVNPTLLFL KVPAQNAIST TFPYTGDPPY SHGTGTGYTM DTVNRTHQYS EKGKWTTNTE TGAPQLNPID GPLPEDNEPS GYAQTDCVL EAMAFLEESH PGIFENSCLE TMEVVQQTRV DRLTQGRQTY DWTLNRNQPA ATALANTIEV FRSNGLTANE S GRLIDFLK DVMESMDKEE MEITTHFQRK RRVRDNMTKK MVTQRTIGKK KQRVNKRSYL IRALTLNTMT KDAERGKLKR RA IATPGMQ IRGFVYFVET LARSICEKLE QSGLPVGGNE KKAKLANVVR KMMTNSQDTE LSFTITGDNT KWNENQNPRM FLA MITYIT KNQPEWFRNV LSIAPIMFSN KMARLGKGYM FESKSMKLRT QIPAEMLASI DLKYFNESTR KKIEKIRPLL IDGT ASLSP GMMMGMFNML STVLGVSILN LGQKRYTKTT YWWDGLQSSD DFALIVNAPN HEGIQAGVDR FYRTCKLVGI NMSKK KSYI NRTGTFEFTS FFYRYGFVAN FSMELPSFGV SGINESADMS IGVTVIKNNM INNDLGPATA QMALQLFIKD YRYTYR CHR GDTQIQTRRS FELKKLWEQT RSKAGLLVSD GGPNLYNIRN LHIPEVCLKW ELMDEDYQGR LCNPLNPFVS HKEIESV NN AVVMPAHGPA KSMEYDAVAT THSWIPKRNR SILNTSQRGI LEDEQMYQKC CNLFEKFFPS SSYRRPVGIS SMVEAMVS R ARIDARIDFE SGRIKKEEFA EIMKICSTIE ELRRQK

UniProtKB: RNA-directed RNA polymerase catalytic subunit

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Macromolecule #3: Polymerase basic protein 2

MacromoleculeName: Polymerase basic protein 2 / type: protein_or_peptide / ID: 3 / Number of copies: 2 / Enantiomer: LEVO
Source (natural)Organism: Influenza A virus (A/nt/60/1968(H3N2)) / Strain: A/nt/60/1968(H3N2)
Molecular weightTheoretical: 89.49593 KDa
Recombinant expressionOrganism: Homo sapiens (human)
SequenceString: MERIKELRNL MSQSRTREIL TKTTVDHMAI IKKYTSGRQE KNPSLRMKWM MAMKYPITAD KRITEMVPER NEQGQTLWSK MSDAGSDRV MVSPLAVTWW NRNGPMTSTV HYPKVYKTYF EKVERLKHGT FGPVHFRNQV KIRRRVDINP GHADLSAKEA Q DVIMEVVF ...String:
MERIKELRNL MSQSRTREIL TKTTVDHMAI IKKYTSGRQE KNPSLRMKWM MAMKYPITAD KRITEMVPER NEQGQTLWSK MSDAGSDRV MVSPLAVTWW NRNGPMTSTV HYPKVYKTYF EKVERLKHGT FGPVHFRNQV KIRRRVDINP GHADLSAKEA Q DVIMEVVF PNEVGARILT SESQLTITKE KKEELQDCKI SPLMVAYMLE RELVRKTRFL PVAGGTSSVY IEVLHLTQGT CW EQMYTPG GEVRNDDVDQ SLIIAARNIV RRAAVSADPL ASLLEMCHST QIGGTRMVDI LRQNPTEEQA VDICKAAMGL RIS SSFSFG GFTFKRTSGS SIKREEELLT GNLQTLKIRV HDGYEEFTMV GKRATAILRK ATRRLVQLIV SGRDEQSVAE AIIV AMVFS QEDCMIKAVR GDLNFVNRAN QRLNPMHQLL RHFQKDAKVL FQNWGIEHID NVMGMIGVLP DMTPSTEMSM RGIRV SKMG VDEYSSTERV VVSIDRFLRV RDQRGNVLLS PEEVSETQGT EKLTITYSSS MMWEINGPES VLVNTYQWII RNWETV KIQ WSQNPTMLYN KMEFEPFQSL VPKAIRGQYS GFVRTLFQQM RDVLGTFDTT QIIKLLPFAA APPKQSRMQF SSLTVNV RG SGMRILVRGN SPAFNYNKTT KRLTILGKDA GTLIEDPDEG TSGVESAVRR GFLILGKEDR RYGPALSINE LSNLAKGE K ANVLIGQGDV VLVMKRKRDS SILTDSQTAT KRIRMAINGW SHPQFEKGGG SGGGSGGSAW SHPQFEKGRS G

UniProtKB: Polymerase basic protein 2

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Macromolecule #4: Nuclear export protein

MacromoleculeName: Nuclear export protein / type: protein_or_peptide / ID: 4 / Number of copies: 2 / Enantiomer: LEVO
Source (natural)Organism: Influenza A virus (strain A/Hong Kong/1/1968 H3N2)
Strain: A/Hong Kong/1/1968 H3N2
Molecular weightTheoretical: 14.355328 KDa
Recombinant expressionOrganism: Homo sapiens (human)
SequenceString:
MDSNTVSSFQ DILLRMSKMQ LGSSSEDLNG MITQFESLKL YRDSLGEAVM RMGDLHSLQN RNGKWREQLG QKFEEIRWLI EEVRHRLKT TENSFEQITF MQALQLLFEV EQEIRTFSFQ LI

UniProtKB: Nuclear export protein

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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

BufferpH: 8
VitrificationCryogen name: ETHANE

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Image recordingFilm or detector model: GATAN K3 (6k x 4k) / Average electron dose: 50.0 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Cs: 0.001 mm / Nominal defocus max: 2.0 µm / Nominal defocus min: 1.0 µm
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Startup modelType of model: INSILICO MODEL
Final reconstructionResolution.type: BY AUTHOR / Resolution: 3.0 Å / Resolution method: FSC 0.143 CUT-OFF / Number images used: 351243
Initial angle assignmentType: MAXIMUM LIKELIHOOD
Final angle assignmentType: MAXIMUM LIKELIHOOD

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