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- EMDB-18822: Structure of avian H5N1 influenza A polymerase in complex with hu... -

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Basic information

Entry
Database: EMDB / ID: EMD-18822
TitleStructure of avian H5N1 influenza A polymerase in complex with human ANP32B.
Map data
Sample
  • Complex: Avian H5N1 apo influenza A polymerase in complex with human ANP32B.
    • Protein or peptide: Acidic leucine-rich nuclear phosphoprotein 32 family member B
    • Protein or peptide: Polymerase acidic protein
    • Protein or peptide: RNA-directed RNA polymerase catalytic subunit
    • Protein or peptide: Polymerase basic protein 2
KeywordsInfluenza virus / replication platform / H5N1 / influenza adaptive mutations / host adaptation / viral polymerase / cryo-EM / HPAI. / VIRAL PROTEIN
Function / homology
Function and homology information


ventricular system development / vasculature development / RNA polymerase binding / roof of mouth development / cap snatching / viral transcription / inner ear development / symbiont-mediated suppression of host mRNA transcription via inhibition of RNA polymerase II activity / negative regulation of cell differentiation / positive regulation of protein export from nucleus ...ventricular system development / vasculature development / RNA polymerase binding / roof of mouth development / cap snatching / viral transcription / inner ear development / symbiont-mediated suppression of host mRNA transcription via inhibition of RNA polymerase II activity / negative regulation of cell differentiation / positive regulation of protein export from nucleus / activation of cysteine-type endopeptidase activity involved in apoptotic process / nucleosome assembly / histone binding / endonuclease activity / regulation of apoptotic process / host cell cytoplasm / RNA-directed RNA polymerase / viral RNA genome replication / nucleotide binding / RNA-dependent RNA polymerase activity / host cell nucleus / RNA binding / extracellular exosome / nucleoplasm / metal ion binding / nucleus / cytoplasm
Similarity search - Function
Acidic leucine-rich nuclear phosphoprotein 32 / U2A'/phosphoprotein 32 family A, C-terminal / occurring C-terminal to leucine-rich repeats / Leucine-rich repeat / Influenza RNA-dependent RNA polymerase subunit PB1 / Influenza RNA-dependent RNA polymerase subunit PB1 / 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 ...Acidic leucine-rich nuclear phosphoprotein 32 / U2A'/phosphoprotein 32 family A, C-terminal / occurring C-terminal to leucine-rich repeats / Leucine-rich repeat / Influenza RNA-dependent RNA polymerase subunit PB1 / Influenza RNA-dependent RNA polymerase subunit PB1 / 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. / Leucine-rich repeat profile. / Leucine-rich repeat / Leucine-rich repeat domain superfamily
Similarity search - Domain/homology
RNA-directed RNA polymerase catalytic subunit / Polymerase acidic protein / Acidic leucine-rich nuclear phosphoprotein 32 family member B
Similarity search - Component
Biological speciesInfluenza A virus / Homo sapiens (human)
Methodsingle particle reconstruction / cryo EM / Resolution: 3.1 Å
AuthorsCarrique L / Staller E / Keown JR / Fan H / Fodor E / Grimes JM
Funding support United Kingdom, 6 items
OrganizationGrant numberCountry
Wellcome Trust200835/Z/16/Z United Kingdom
Wellcome Trust222510/Z/21/Z United Kingdom
Medical Research Council (MRC, United Kingdom)MR/R009945/1 United Kingdom
Wellcome Trust093305/Z/10/Z United Kingdom
Wellcome Trust060208/Z/00/Z United Kingdom
Wellcome Trust203141/Z/16/Z United Kingdom
CitationJournal: Nat Commun / Year: 2024
Title: Structures of H5N1 influenza polymerase with ANP32B reveal mechanisms of genome replication and host adaptation.
Authors: Ecco Staller / Loïc Carrique / Olivia C Swann / Haitian Fan / Jeremy R Keown / Carol M Sheppard / Wendy S Barclay / Jonathan M Grimes / Ervin Fodor /
Abstract: Avian influenza A viruses (IAVs) pose a public health threat, as they are capable of triggering pandemics by crossing species barriers. Replication of avian IAVs in mammalian cells is hindered by ...Avian influenza A viruses (IAVs) pose a public health threat, as they are capable of triggering pandemics by crossing species barriers. Replication of avian IAVs in mammalian cells is hindered by species-specific variation in acidic nuclear phosphoprotein 32 (ANP32) proteins, which are essential for viral RNA genome replication. Adaptive mutations enable the IAV RNA polymerase (FluPolA) to surmount this barrier. Here, we present cryo-electron microscopy structures of monomeric and dimeric avian H5N1 FluPolA with human ANP32B. ANP32B interacts with the PA subunit of FluPolA in the monomeric form, at the site used for its docking onto the C-terminal domain of host RNA polymerase II during viral transcription. ANP32B acts as a chaperone, guiding FluPolA towards a ribonucleoprotein-associated FluPolA to form an asymmetric dimer-the replication platform for the viral genome. These findings offer insights into the molecular mechanisms governing IAV genome replication, while enhancing our understanding of the molecular processes underpinning mammalian adaptations in avian-origin FluPolA.
History
DepositionNov 2, 2023-
Header (metadata) releaseMay 8, 2024-
Map releaseMay 8, 2024-
UpdateMay 29, 2024-
Current statusMay 29, 2024Processing site: PDBe / Status: Released

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Structure visualization

Supplemental images

emd_18822.png

Downloads & links

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Map

FileDownload / File: emd_18822.map.gz / Format: CCP4 / Size: 103 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Voxel sizeX=Y=Z: 0.932 Å
Density
Contour LevelBy AUTHOR: 0.248
Minimum - Maximum-1.6138949 - 2.0128672
Average (Standard dev.)0.00018717484 (±0.04727994)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions300300300
Spacing300300300
CellA=B=C: 279.6 Å
α=β=γ: 90.0 °

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Supplemental data

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Mask #1

Fileemd_18822_msk_1.map
Projections & Slices
AxesZYX

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Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: #1

Fileemd_18822_half_map_1.map
Projections & Slices
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Histogram

Histogram (log scale)

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Half map: #2

Fileemd_18822_half_map_2.map
Projections & Slices
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Sample components

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Entire : Avian H5N1 apo influenza A polymerase in complex with human ANP32B.

EntireName: Avian H5N1 apo influenza A polymerase in complex with human ANP32B.
Components
  • Complex: Avian H5N1 apo influenza A polymerase in complex with human ANP32B.
    • Protein or peptide: Acidic leucine-rich nuclear phosphoprotein 32 family member B
    • Protein or peptide: Polymerase acidic protein
    • Protein or peptide: RNA-directed RNA polymerase catalytic subunit
    • Protein or peptide: Polymerase basic protein 2

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Supramolecule #1: Avian H5N1 apo influenza A polymerase in complex with human ANP32B.

SupramoleculeName: Avian H5N1 apo influenza A polymerase in complex with human ANP32B.
type: complex / ID: 1 / Parent: 0 / Macromolecule list: all
Source (natural)Organism: Influenza A virus / Strain: A/turkey/Turkey/1/2005(H5N1)

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Macromolecule #1: Acidic leucine-rich nuclear phosphoprotein 32 family member B

MacromoleculeName: Acidic leucine-rich nuclear phosphoprotein 32 family member B
type: protein_or_peptide / ID: 1 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Homo sapiens (human)
Molecular weightTheoretical: 28.816652 KDa
Recombinant expressionOrganism: Spodoptera frugiperda (fall armyworm)
SequenceString: MDMKRRIHLE LRNRTPAAVR ELVLDNCKSN DGKIEGLTAE FVNLEFLSLI NVGLISVSNL PKLPKLKKLE LSENRIFGGL DMLAEKLPN LTHLNLSGNK LKDISTLEPL KKLECLKSLD LFNCEVTNLN DYRESVFKLL PQLTYLDGYD REDQEAPDSD A EVDGVDEE ...String:
MDMKRRIHLE LRNRTPAAVR ELVLDNCKSN DGKIEGLTAE FVNLEFLSLI NVGLISVSNL PKLPKLKKLE LSENRIFGGL DMLAEKLPN LTHLNLSGNK LKDISTLEPL KKLECLKSLD LFNCEVTNLN DYRESVFKLL PQLTYLDGYD REDQEAPDSD A EVDGVDEE EEDEEGEDEE DEDDEDGEEE EFDEEDDEDE DVEGDEDDDE VSEEEEEFGL DEEDEDEDED EEEEEGGKGE KR KRETDDE GEDD

UniProtKB: Acidic leucine-rich nuclear phosphoprotein 32 family member B

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Macromolecule #2: Polymerase acidic protein

MacromoleculeName: Polymerase acidic protein / type: protein_or_peptide / ID: 2 / Details: Polymerase Acidic Protein A / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Influenza A virus / Strain: Influenza A virus (A/turkey/Turkey/1/2005(H5N1))
Molecular weightTheoretical: 82.600281 KDa
Recombinant expressionOrganism: Spodoptera frugiperda (fall armyworm)
SequenceString: MEDFVRQCFN PMIVELAEKA MKEYGEDPKI ETNKFAAICT HLEVCFMYSD FHFIDERSES IIVESGDPNA LLKHRFEIIE GRDRTMAWT VVNSICNTTG VEKPKFLPDL YDYKENRFIE IGVTRREVHT YYLEKANKIK SEKTHIHIFS FTGEEMATKA D YTLDEESR ...String:
MEDFVRQCFN PMIVELAEKA MKEYGEDPKI ETNKFAAICT HLEVCFMYSD FHFIDERSES IIVESGDPNA LLKHRFEIIE GRDRTMAWT VVNSICNTTG VEKPKFLPDL YDYKENRFIE IGVTRREVHT YYLEKANKIK SEKTHIHIFS FTGEEMATKA D YTLDEESR ARIKTRLFTI RQEMATRGLW DSFRQSERGE ETIEEKFEIT GTMRRLADQS LPPNFSSLEN FRAYVDGFEP NG CIEGKLS QMSKEVNARI EPFLKTTPRP LKLPDGPPCS QRSKFLLMDA LKLSIEDPSH EGEGIPLYDA IKCMKTFFGW KEP NIVKPH EKGINPNYLL TWKQVLAELQ DIENEEKIPK TKNMKKTSQL RWALGENMAP EKVDFEDCKD VSDLKQYDSD EPES RSLAS WIQSEFNKAC ELTDSSWIEL DEIGEDVAPI EHIASMRRNY FTAEVSHCRA TEYIMKGVYI NTALLNASCA AMDDF QLIP MISKCRTKEG RRKTNLYGFI IKGRSHLRND TDVVNFVSME FSLTDPRLEP HKWEKYCVLE IGNMLLRTAV GRVSRP MFL YVRTNGTSKI KMKWGMEMRR CLLQSLQQIE SMIEAESSVK EKDMTKEFFE NKSETWPIGE SPKGVEEGSI GKVCRTL LA KSVFNSLYAS SQLEGFSAES RKLLLIAQAL RDNLEPGTFD LGGLYEAIEE CLINDPWVLL NASWFNSFLA HALK

UniProtKB: Polymerase acidic protein

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Macromolecule #3: RNA-directed RNA polymerase catalytic subunit

MacromoleculeName: RNA-directed RNA polymerase catalytic subunit / type: protein_or_peptide / ID: 3 / Number of copies: 1 / Enantiomer: LEVO / EC number: RNA-directed RNA polymerase
Source (natural)Organism: Influenza A virus / Strain: Influenza A virus (A/turkey/Turkey/1/2005(H5N1))
Molecular weightTheoretical: 86.616312 KDa
Recombinant expressionOrganism: Spodoptera frugiperda (fall armyworm)
SequenceString: MDVNPTLLFL KVPVQNAIST TFPYTGDPPY SHGTGTGYTM DTVNRTHQYS EKGKWTKNTE TGAPQLNPID GPLPEDNEPS GYAQTDCVL EAMAFLEESH PGIFENSCLE TMEIVQQTRV DKLTQGRQTY DWTLNRNQPA ATALANTIEI FRSNGLTANE S GRLIDFLK ...String:
MDVNPTLLFL KVPVQNAIST TFPYTGDPPY SHGTGTGYTM DTVNRTHQYS EKGKWTKNTE TGAPQLNPID GPLPEDNEPS GYAQTDCVL EAMAFLEESH PGIFENSCLE TMEIVQQTRV DKLTQGRQTY DWTLNRNQPA ATALANTIEI FRSNGLTANE S GRLIDFLK DVMESMDKEE MEITTHFQRK RRVRDNMTKK MVTQRTIGKK KQRLNKKSYL IRALTLNTMT KDAERGKLKR RA IATPGMQ IRGFVYFVET LARSICEKLE QSGLPVGGNE KKAKLANVVR KMMTNSQDTE LSFTITGDNT KWNENQNPRM FLA MITYIT RNQPEWFRNV LSIAPIMFSN KMARLGRGYM FESKSMKLRT QIPAEMLANI DLKYFNELTK KKIEKIRPLL IDGT ASLSP GMMMGMFNML STVLGVSILN LGQKRYTKTT YWWDGLQSSD DFALIVNAPN HEGIQAGVDR FYRTCKLVGI NMSKK KSYI NRTGTFEFTS FFYRYGFVAN FSMELPSFGV SGINESADMS IGVTVIKNNM INNDLGPATA QMALQLFIKD YRYTYR CHR GDTQIQTRRS FELEKLWEQT RSKAGLLVSD GGPNLYNIRN LHIPEVCLKW ELMDEDYQGR LCNPLNPFVS HKEIESV NN AVVMPAHGPA KSMEYDAVAT THSWIPKRNR SILNTSQRGI LEDEQMYQKC CNLFEKFFPS SSYRRPVGIS SMVEAMVS R ARIDARIDFE SGRIKKEEFA EIMKICSTIE ELRRPK

UniProtKB: RNA-directed RNA polymerase catalytic subunit

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Macromolecule #4: Polymerase basic protein 2

MacromoleculeName: Polymerase basic protein 2 / type: protein_or_peptide / ID: 4 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Influenza A virus / Strain: Influenza A virus (A/turkey/Turkey/1/2005(H5N1))
Molecular weightTheoretical: 101.380016 KDa
Recombinant expressionOrganism: Spodoptera frugiperda (fall armyworm)
SequenceString: MERIKELRDL MSQSRTREIL TKTTVDHMAI IKKYTSGRQE KNPALRMKWM MAMKYPITAD KRIIEMIPER NEQGQTLWSK TNDAGSDRV MVSPLAVTWW NRNGPTTSTV HYPKVYKTYF EKVERLKHGT FGPVHFRNQV KIRRRVDTNP GHADLSAKEA Q DVIMEVVF ...String:
MERIKELRDL MSQSRTREIL TKTTVDHMAI IKKYTSGRQE KNPALRMKWM MAMKYPITAD KRIIEMIPER NEQGQTLWSK TNDAGSDRV MVSPLAVTWW NRNGPTTSTV HYPKVYKTYF EKVERLKHGT FGPVHFRNQV KIRRRVDTNP GHADLSAKEA Q DVIMEVVF PNEVGARILT SESQLTITKE KKEELQDCKI APLMVAYMLE RELVRKTRFL PVAGGTSSVY IEVLHLTQGT CW EQMYTPG GEVRNDDVDQ SLIIAARNIV RRATVSADPL ASLLEMCHST QIGGIRMVDI LRQNPTEEQA VDICKAAMGL RIS SSFSFG GFTFKRTSGS SVTKEEEVLT GNLQTLKIRV HEGYEEFTMV GQRATAILRK ATRRLIQLIV SGRNEQSIAE AIIV AMVFS QEDCMIKAVR GDLNFVNRAN QRLNPMHQLL RHFQKDAKVL FQNWGTEPID NVMGMIGILP DMTPSTEMSL RGVRV SKMG VDEYSSTERV VVSIDRFLRV RDQRGNVLLS PEEVSETQGT EKLTITYSSS MMWEINGPES VLVNTYQWII RNWETV KIQ WSQDPTMLYN KMEFEPFQSL VPKAARGQYS GFVRTLFQQM RDVLGTFDTV QIIKLLPFAA APPKQSRMQF SSLTVNV RG SGMRILIRGN SPVFNYNKAT KRLTVLGKDA GALTEDPDEG TAGVESAVLR GFLILGKEDK RYGPALSINE LSNLAKGE K ANVLIGQGDV VLVMKRKRDS SILTDSQTAT KRIRMAINEL KTAALAQHDE AVDNKFNKEQ QNAFYEILHL PNLNEEQRN AFIQSLKDDP SQSANLLAEA KKLNDAQAPK VDNKFNKEQQ NAFYEILHLP NLNEEQRNAF IQSLKADPSQ SANLLAEAKK LNGAQAPKV DANSAGKST

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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

Concentration0.3 mg/mL
BufferpH: 7.5
Component:
ConcentrationName
25.0 mMHEPES
150.0 mMKCl
5.0 %Glycerol
GridModel: Quantifoil R2/1 / Material: COPPER / Mesh: 200 / Support film - Material: CARBON / Support film - topology: HOLEY
VitrificationCryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 293 K / Instrument: FEI VITROBOT MARK IV

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Electron microscopy

MicroscopeTFS KRIOS
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsC2 aperture diameter: 50.0 µm / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELDBright-field microscopy / Cs: 2.7 mm / Nominal defocus max: 24.0 µm / Nominal defocus min: 5.0 µm
Specialist opticsEnergy filter - Name: TFS Selectris X / Energy filter - Slit width: 10 eV
Sample stageSpecimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Cooling holder cryogen: NITROGEN
Image recordingFilm or detector model: TFS FALCON 4i (4k x 4k) / Average electron dose: 50.0 e/Å2
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Startup modelType of model: INSILICO MODEL
Initial angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: cryoSPARC
Final 3D classificationNumber classes: 10 / Avg.num./class: 25000 / Software - Name: cryoSPARC (ver. 4.2)
Final angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: cryoSPARC (ver. 4.2)
Final reconstructionNumber classes used: 1 / Applied symmetry - Point group: C1 (asymmetric) / Resolution.type: BY AUTHOR / Resolution: 3.1 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: cryoSPARC (ver. V4.2) / Number images used: 63000
FSC plot (resolution estimation)

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Atomic model buiding 1

Initial model
ChainDetailsPDB ID
source_name: AlphaFold, initial_model_type: in silico modelANP32 model
source_name: PDB, initial_model_type: experimental model

6rr7

RefinementSpace: REAL / Protocol: RIGID BODY FIT
Output model

PDB-8r1l:
Structure of avian H5N1 influenza A polymerase in complex with human ANP32B.

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