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- EMDB-33986: Cryo-EM structure of RNA polymerase in complex with P protein tet... -

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Basic information

Entry
Database: EMDB / ID: EMD-33986
TitleCryo-EM structure of RNA polymerase in complex with P protein tetramer of Newcastle disease virus
Map data
Sample
  • Complex: NDV L-P complex
    • Protein or peptide: NDV P protein
    • Protein or peptide: RNA-directed RNA polymerase L
Keywordscryo-EM / L-P complex / Newcastle disease virus / VIRUS
Function / homology
Function and homology information


GDP polyribonucleotidyltransferase / Hydrolases; Acting on acid anhydrides; In phosphorus-containing anhydrides / Transferases; Transferring one-carbon groups; Methyltransferases / virion component / mRNA 5'-cap (guanine-N7-)-methyltransferase activity / host cell cytoplasm / RNA-directed RNA polymerase / RNA-dependent RNA polymerase activity / GTPase activity / ATP binding / cytoplasm
Similarity search - Function
RNA-directed RNA polymerase, paramyxovirus / Phosphoprotein P soyouz module / N-terminal region of Paramyxovirinae phosphoprotein (P) / P/V phosphoprotein, paramyxoviral / Paramyxovirus P/V phosphoprotein C-terminal / Mononegavirales RNA-directed RNA polymerase catalytic domain / Mononegavirus L protein 2-O-ribose methyltransferase / Mononegavirales mRNA-capping domain V / RNA-directed RNA polymerase L, C-terminal / Mononegavirales RNA dependent RNA polymerase ...RNA-directed RNA polymerase, paramyxovirus / Phosphoprotein P soyouz module / N-terminal region of Paramyxovirinae phosphoprotein (P) / P/V phosphoprotein, paramyxoviral / Paramyxovirus P/V phosphoprotein C-terminal / Mononegavirales RNA-directed RNA polymerase catalytic domain / Mononegavirus L protein 2-O-ribose methyltransferase / Mononegavirales mRNA-capping domain V / RNA-directed RNA polymerase L, C-terminal / Mononegavirales RNA dependent RNA polymerase / Mononegavirales mRNA-capping region V / RdRp of negative ssRNA viruses with non-segmented genomes catalytic domain profile. / Mononegavirus L protein 2'-O-ribose methyltransferase domain profile.
Similarity search - Domain/homology
RNA-directed RNA polymerase L / Phosphoprotein
Similarity search - Component
Biological speciesAvian orthoavulavirus 1
Methodsingle particle reconstruction / cryo EM / Resolution: 3.0 Å
AuthorsChen Y / Jingyuan C
Funding support China, 1 items
OrganizationGrant numberCountry
National Basic Research Program of China (973 Program)2017YFC0840300 China
CitationJournal: Nat Commun / Year: 2023
Title: Structure of the Newcastle Disease Virus L protein in complex with tetrameric phosphoprotein.
Authors: Jingyuan Cong / Xiaoying Feng / Huiling Kang / Wangjun Fu / Lei Wang / Chenlong Wang / Xuemei Li / Yutao Chen / Zihe Rao /
Abstract: Newcastle disease virus (NDV) belongs to Paramyxoviridae, which contains lethal human and animal pathogens. NDV RNA genome is replicated and transcribed by a multifunctional 250 kDa RNA-dependent ...Newcastle disease virus (NDV) belongs to Paramyxoviridae, which contains lethal human and animal pathogens. NDV RNA genome is replicated and transcribed by a multifunctional 250 kDa RNA-dependent RNA polymerase (L protein). To date, high-resolution structure of NDV L protein complexed with P protein remains to be elucidated, limiting our understanding of the molecular mechanisms of Paramyxoviridae replication/transcription. Here, we used cryo-EM and enzymatic assays to investigate the structure-function relationship of L-P complex. We found that C-terminal of CD-MTase-CTD module of the atomic-resolution L-P complex conformationally rearranges, and the priming/intrusion loops are likely in RNA elongation conformations different from previous structures. The P protein adopts a unique tetrameric organization and interacts with L protein. Our findings indicate that NDV L-P complex represents elongation state distinct from previous structures. Our work greatly advances the understanding of Paramyxoviridae RNA synthesis, revealing how initiation/elongation alternates, providing clues for identifying therapeutic targets against Paramyxoviridae.
History
DepositionAug 2, 2022-
Header (metadata) releaseFeb 15, 2023-
Map releaseFeb 15, 2023-
UpdateAug 30, 2023-
Current statusAug 30, 2023Processing site: PDBj / Status: Released

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Structure visualization

Supplemental images

emd_33986.png

Downloads & links

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Map

FileDownload / File: emd_33986.map.gz / Format: CCP4 / Size: 30.5 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Voxel sizeX=Y=Z: 1.04 Å
Density
Contour LevelBy AUTHOR: 0.035
Minimum - Maximum-0.12501945 - 0.21997634
Average (Standard dev.)-0.000022567328 (±0.0076793334)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions200200200
Spacing200200200
CellA=B=C: 208.0 Å
α=β=γ: 90.0 °

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Supplemental data

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Half map: #1

Fileemd_33986_half_map_1.map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: #2

Fileemd_33986_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 : NDV L-P complex

EntireName: NDV L-P complex
Components
  • Complex: NDV L-P complex
    • Protein or peptide: NDV P protein
    • Protein or peptide: RNA-directed RNA polymerase L

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Supramolecule #1: NDV L-P complex

SupramoleculeName: NDV L-P complex / type: complex / ID: 1 / Parent: 0 / Macromolecule list: all
Source (natural)Organism: Avian orthoavulavirus 1

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Macromolecule #1: NDV P protein

MacromoleculeName: NDV P protein / type: protein_or_peptide / ID: 1 / Number of copies: 4 / Enantiomer: LEVO
Source (natural)Organism: Avian orthoavulavirus 1
Molecular weightTheoretical: 41.738379 KDa
Recombinant expressionOrganism: Spodoptera frugiperda (fall armyworm)
SequenceString: MATFTDAEID ELFETSGTVI DSIITAQGKP VETVGRSAIP QGKTKALSLA WEKHGNTNTP AAQESAGEQD QHGQNQASNS NRATPEEGP HSSQAQAATQ PQEDANESQL KTGASSSLLS MLDKLSNKSS NAKKGPPQSP PQQALHSKGS PAVEQTQHGA N QGRAQQET ...String:
MATFTDAEID ELFETSGTVI DSIITAQGKP VETVGRSAIP QGKTKALSLA WEKHGNTNTP AAQESAGEQD QHGQNQASNS NRATPEEGP HSSQAQAATQ PQEDANESQL KTGASSSLLS MLDKLSNKSS NAKKGPPQSP PQQALHSKGS PAVEQTQHGA N QGRAQQET GHQAAPSPGP PGTGVNIAFP GQRGVSPQSV GATQPAPQSG QNQGSTPASA DHVQPPVDFV QAMMSMMEAI SQ RVSKIDY QLDLVLKQTS SIPTMRSEIQ QLKTSVAVME ANLGMMKILD PGCANVSSLS DLRAVAKSHP VLIAGPGDPS PYV TQGGEI ALNKLSQPVP HPSDLIKHAT SGGPDIGIER DTVRALILSR PMHPSSSSKL LSKLDSAGSV EEIRKIKRLA LNG

UniProtKB: Phosphoprotein

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Macromolecule #2: RNA-directed RNA polymerase L

MacromoleculeName: RNA-directed RNA polymerase L / type: protein_or_peptide / ID: 2 / Number of copies: 1 / Enantiomer: LEVO / EC number: RNA-directed RNA polymerase
Source (natural)Organism: Avian orthoavulavirus 1
Molecular weightTheoretical: 249.183188 KDa
Recombinant expressionOrganism: Spodoptera frugiperda (fall armyworm)
SequenceString: MAGSGSERAE HQIILPESHL SSPLVKHKLL YYWKLTGLPL PDECDFDHLI LSRQWKKILE SSTPDIERMI KLGRSVHQTL SHSSKLTGI LHPRCLEDLV GLDIPDSTNK FRRIEKKIQI HNTRYGEPFT RLCSYVEKKL LGSSWTHKIR RSEEFDSLRT D PAFWFHSS ...String:
MAGSGSERAE HQIILPESHL SSPLVKHKLL YYWKLTGLPL PDECDFDHLI LSRQWKKILE SSTPDIERMI KLGRSVHQTL SHSSKLTGI LHPRCLEDLV GLDIPDSTNK FRRIEKKIQI HNTRYGEPFT RLCSYVEKKL LGSSWTHKIR RSEEFDSLRT D PAFWFHSS WSTAKFAWLH VKQIQRHLIV AARTRSASNK LVTLSHRSGQ VFITPELVIV THTNENKFTC LSQELVLMYA DM MEGRDMV NIISSTAVHL RCLAEKIDDI LRLVDALARD LGNQVYDVVA LMEGFAYGAV QLLEPSGTFA GDFFSFNLQE LRD TLICLL PQRIADSVTH AIANIFSGLE QNQAAEMLCL LRLWGHPLLE SRAAAKAVRA QMCAPKMVDF DMILQVLSFF KGTI INGYR KKNAGVWPRV KAHTIYGNVI AQLHADSAEI SHDIMLREYK NLSAIEFEAC IEYDPVTNLS MFLKDKAIAH PRNNW LASF RRNLLSEEQK KNVQDSTSTN RLLIEFLESN DFDPYKEMEY LTTLEYLRDD SVAVSYSLKE EEVKVNGRIF AKLTKK LRN CQVMAEGILA DQIAPFFQGN GVIQDSISLT KSMLAMSQLS YNSNRKRITD CKERVSSSRN HDLKGKHRRR VATFITT DL QKYCLNWRYQ TIKLFAHAIN QLMGLPHFFE WIHLRLMDTT MFVGDPFNPP SDPTDYDLTK VPNDDIYIVS ARGGIEGL C QKLWTMISIA AIQLAAARSH CRVACMVQGD NQVIAVTREV RPDDSPESVL TQLHEASDNF FRELIHVNHL IGHNLKDRE TIRSDTFFIY SKRIFKDGAI LSQVLKNSSK LVLVSGDLSE NTVMSCANIS STVARLCENG LPKDFCYYLN YLMSCIQTYF DSEFSITSS TQSGSNQSWI NDIPFIHSYV LTPAQLGGLS NLQYSRLYTR NIGDPGTTAF AEVKRLEAVG LLGPNIMTNI L TRPPGNGD WASLCNDPYS FNFESVASPS IVLKKHTQRV LFETCSNPLL SGVHTEDNEA EEKALAEYLL NQEVIHPRVA HA IMEASSV GRRKQIQGLV DTTNTVIKIA LSRKPLGIKR LARIINYSSM HAMLFRDDVF LSNRANHPLV SSDMCSLALA DYA RNRSWS PLTGGRKILG VSNPDTIELV EGEILSISGG CSKCDSGDEQ FTWFHLPSNI ELTDDTSKNP PMRVPYLGSK TQER RAASL AKIAHMSPHV KAALRASSVL IWAYGDNDIN WTAALKLARS RCNISSEYLR LLSPLPTAGN LQHRLDDGIT QMTFT PASL YRVSPYVHIS NDSQRLFTEE GVKEGNVVYQ QIMLLGLSLI ESLFPMTVTK TYDEITLHLH SKFSCCIREA PVAVPF ELT GVAPDLRVVA SNKFMYDPNP VAEGDFARLD LAIFKSYELN LESYSTVELM NILSISSGKL IGQSVVSYDE ETSIKND AI IVYDNTRNWI SEAQNSDVVR LFEYAALEVL LDCSYQLYYL RVRGLNNVVL YMSDLYKNMP GILLSNIAAT ISHPIIHS R LHTVGLISHD GSHQLADTDF IELSAKLLVS CTRRVVSGLY AGNKYDLLFP SVLDDNLNEK MLQLISRLCC LYTVLFATT REIPKIRGLP AEEKCAMLTE YLLSDAVRPL LSPEQVDSIT SPSIVTFPAN LYYMSRKSLN LIREREDRDS ILALMFPQEP LFEFPLVQD IGARVKDQLT MKPAAFLHEL DLSAPARYDA YTLEQARSDC ALADMGEDQL VRYLFRGVGT ASSSWYKASH L LSVPEIRC ARHGNSLYLA EGSGAIMSLL ELHIPHETIY YNTLFSNEMN PPQRHFGPTP TQFLNSVVYR NLQAEVPCKD GF VQEFRTL WRENTEESDL TSDKAVGYIT SVVPYRSVSL LHCDIEIPPG SNQSLLDQLA TNLSLIAMHS VREGGVVIVK ILY SMGYYF HLLVNLFTPC SVKGYVLSNG YACRGDMECY VVFVMGYLGG PTFVNEVVRM AKTLIQRHGT LLAKSDETAL MALF TSQKQ RVDNILSSPL PRLAKLLRRN IDTALIEAGG QPVRPFCAES LVNTLSDITQ TTQVIASHID TVIRSVIYME AEGDL ADTV FLFTPYNLSI DGKKRTSLKQ CTRQILEVTI LGLGPEDLNR VGDIISLILR GTISLEDLIP LRTYLKMSTC PKYLKS VLG LTKLREMFSD GSMLYLTRAQ QKFYMKTVGN AVKGYYNSSK NENLYFQG

UniProtKB: RNA-directed RNA polymerase L

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Experimental details

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Structure determination

Methodcryo EM
Processingsingle particle reconstruction
Aggregation state3D array

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Sample preparation

BufferpH: 8
VitrificationCryogen name: ETHANE

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELDBright-field microscopy / Nominal defocus max: 2.5 µm / Nominal defocus min: 1.2 µm
Image recordingFilm or detector model: GATAN K2 SUMMIT (4k x 4k) / Average electron dose: 60.0 e/Å2
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Startup modelType of model: NONE
Initial angle assignmentType: ANGULAR RECONSTITUTION
Final angle assignmentType: ANGULAR RECONSTITUTION
Final reconstructionResolution.type: BY AUTHOR / Resolution: 3.0 Å / Resolution method: FSC 0.143 CUT-OFF / Number images used: 455000

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