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- EMDB-30387: Structure of Lassa virus polymerase bound to Z matrix protein -

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Basic information

Entry
Database: EMDB / ID: EMD-30387
TitleStructure of Lassa virus polymerase bound to Z matrix protein
Map data
Sample
  • Complex: Lassa virus polymerase in complex with Z matrix protein
    • Protein or peptide: RNA-directed RNA polymerase L
    • Protein or peptide: RING finger protein Z
  • Ligand: MANGANESE (II) ION
  • Ligand: ZINC ION
Function / homology
Function and homology information


negative stranded viral RNA replication / cap snatching / viral budding via host ESCRT complex / viral budding from plasma membrane / virion component / host cell cytoplasm / Hydrolases; Acting on ester bonds / host cell perinuclear region of cytoplasm / hydrolase activity / RNA-directed RNA polymerase ...negative stranded viral RNA replication / cap snatching / viral budding via host ESCRT complex / viral budding from plasma membrane / virion component / host cell cytoplasm / Hydrolases; Acting on ester bonds / host cell perinuclear region of cytoplasm / hydrolase activity / RNA-directed RNA polymerase / RNA-dependent RNA polymerase activity / nucleotide binding / host cell plasma membrane / RNA binding / zinc ion binding / membrane / metal ion binding
Similarity search - Function
RING finger protein Z, zinc finger / RING finger protein Z, arenaviridae / Protein Z, RING-type zinc finger superfamily / P-11 zinc finger / RNA polymerase, arenaviral / RNA endonuclease, cap-snatching / Arenavirus RNA polymerase / Arenavirus cap snatching domain / : / RNA-directed RNA polymerase, negative-strand RNA virus / RdRp of negative ssRNA viruses with segmented genomes catalytic domain profile.
Similarity search - Domain/homology
RING finger protein Z / RNA-directed RNA polymerase L
Similarity search - Component
Biological speciesLassa mammarenavirus
Methodsingle particle reconstruction / cryo EM / Resolution: 3.88 Å
AuthorsXu X / Peng R / Peng Q / Shi Y
CitationJournal: Nat Microbiol / Year: 2021
Title: Cryo-EM structures of Lassa and Machupo virus polymerases complexed with cognate regulatory Z proteins identify targets for antivirals.
Authors: Xin Xu / Ruchao Peng / Qi Peng / Min Wang / Ying Xu / Sheng Liu / Xiaolin Tian / Haiteng Deng / Yimin Tong / Xiaoyou Hu / Jin Zhong / Peiyi Wang / Jianxun Qi / George F Gao / Yi Shi /
Abstract: Zoonotic arenaviruses can lead to life-threating diseases in humans. These viruses encode a large (L) polymerase that transcribes and replicates the viral genome. At the late stage of replication, ...Zoonotic arenaviruses can lead to life-threating diseases in humans. These viruses encode a large (L) polymerase that transcribes and replicates the viral genome. At the late stage of replication, the multifunctional Z protein interacts with the L polymerase to shut down RNA synthesis and initiate virion assembly. However, the mechanism by which the Z protein regulates the activity of L polymerase is unclear. Here, we used cryo-electron microscopy to resolve the structures of both Lassa and Machupo virus L polymerases in complex with their cognate Z proteins, and viral RNA, to 3.1-3.9 Å resolutions. These structures reveal that Z protein binding induces conformational changes in two catalytic motifs of the L polymerase, and restrains their conformational dynamics to inhibit RNA synthesis, which is supported by hydrogen-deuterium exchange mass spectrometry analysis. Importantly, we show, by in vitro polymerase reactions, that Z proteins of Lassa and Machupo viruses can cross-inhibit their L polymerases, albeit with decreased inhibition efficiencies. This cross-reactivity results from a highly conserved determinant motif at the contacting interface, but is affected by other variable auxiliary motifs due to the divergent evolution of Old World and New World arenaviruses. These findings could provide promising targets for developing broad-spectrum antiviral drugs.
History
DepositionJul 17, 2020-
Header (metadata) releaseMay 5, 2021-
Map releaseMay 5, 2021-
UpdateNov 17, 2021-
Current statusNov 17, 2021Processing site: PDBj / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 0.02
  • Imaged by UCSF Chimera
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  • Surface view colored by radius
  • Surface level: 0.02
  • Imaged by UCSF Chimera
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  • Surface view with fitted model
  • Atomic models: PDB-7ckl
  • Surface level: 0.03
  • Imaged by UCSF Chimera
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Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

emd_30387.png

Downloads & links

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Map

FileDownload / File: emd_30387.map.gz / Format: CCP4 / Size: 22.2 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Voxel sizeX=Y=Z: 1.36 Å
Density
Contour LevelBy AUTHOR: 0.02 / Movie #1: 0.02
Minimum - Maximum-0.18290497 - 0.2994877
Average (Standard dev.)0.00048843806 (±0.0074925823)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions180180180
Spacing180180180
CellA=B=C: 244.8 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z1.361.361.36
M x/y/z180180180
origin x/y/z0.0000.0000.000
length x/y/z244.800244.800244.800
α/β/γ90.00090.00090.000
MAP C/R/S123
start NC/NR/NS000
NC/NR/NS180180180
D min/max/mean-0.1830.2990.000

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Supplemental data

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Sample components

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Entire : Lassa virus polymerase in complex with Z matrix protein

EntireName: Lassa virus polymerase in complex with Z matrix protein
Components
  • Complex: Lassa virus polymerase in complex with Z matrix protein
    • Protein or peptide: RNA-directed RNA polymerase L
    • Protein or peptide: RING finger protein Z
  • Ligand: MANGANESE (II) ION
  • Ligand: ZINC ION

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Supramolecule #1: Lassa virus polymerase in complex with Z matrix protein

SupramoleculeName: Lassa virus polymerase in complex with Z matrix protein
type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1-#2
Source (natural)Organism: Lassa mammarenavirus
Recombinant expressionOrganism: Spodoptera frugiperda (fall armyworm)
Molecular weightExperimental: 260 KDa

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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 / EC number: RNA-directed RNA polymerase
Source (natural)Organism: Lassa mammarenavirus
Molecular weightTheoretical: 253.505828 KDa
Recombinant expressionOrganism: Spodoptera frugiperda (fall armyworm)
SequenceString: MEEDIACVKD LVSKYLANNE RLSRQKLAFL VQTEPRMLLM EGLKLLSLCI EVDSCNANGC EHNSEDKSVE RILHDHGVLT PSLCFVVPD GYKLTGNVLI LLECFVRSSP ANFEQKYVED FKKLEQLKED LKSVDINLIP LIDGRTSFYN EQIPDWVNDK L RDTLFSLL ...String:
MEEDIACVKD LVSKYLANNE RLSRQKLAFL VQTEPRMLLM EGLKLLSLCI EVDSCNANGC EHNSEDKSVE RILHDHGVLT PSLCFVVPD GYKLTGNVLI LLECFVRSSP ANFEQKYVED FKKLEQLKED LKSVDINLIP LIDGRTSFYN EQIPDWVNDK L RDTLFSLL KYAQESNSLF EESEYSRLCE SLSMTSGRLS GVESLNALLD NRSNHYEEVI ASCHQGINNK LTAHEVKLQI EE EYQVFRN RLRKGEIEGQ FLKVEKNQLL NEFNNLYADK VAEKDSVEHL THQFKRASPI LRFLYANISK GDNGEGNLII GEC QMQCWR SFLNKVKSMR ILNTRRKLLL IFDALILLAS KHDQVRKKPL RGWLGTCFVS VNDRLVSLES TKKDLKKWVE RRQQ VERSR TMQSFQCPSK NQILNSIFQK TISKATTALR DVGISVDHYK IDMEVICPDG YDLIMDFDVS GVTPTISYQR SEEEA FPYI MGDVDLLKTT DLERLSSLSL ALVNSMKTSS TVKLRQNEFG PARYQVVKCK EAYCQEFSLG ETEFQLIYQK TGECSK CYA INDNRVGEVC SFYADPKRYF PAIFSAEVLQ TTVSTMISWI EDCNELEEQL DKIRSLTKMI LILILAHPSK RSQKLLQ NL RYFIMAYVSD YYHKDLIDKV REELITDVEF LLYRLLRTLM GLVLSEDVKS MMTNRFKFIL NISYMCHFIT KETPDRLT D QIKCFEKFLE PKVKFGHVSI NPADTATEEE LDDMVYNAKK FLSKGGCTSA KGPSYKKPGV SKKYLSLLTS SFNNGSLFK EREVKKEIKD PLITSGCATA LDLASNKSVV VNKYTDGSRV LNYDFNKLTA LAVSQLTEVF SRKGKHLLNK QDYEYKVQQA MSNLVLGSK QHKGDADEAD LDEILLDGGA STYFNQLKET VEKIVDQYRE PVKMGSGSND GDQPSINDLD EIVSNKFYIR L IKGELSNH MVEDFDHDVL PDKFYEEFCD AVYENSKLKE KYFYCGHMSQ CPIGELTKAV STRTYLDHEY FQCFKSILLI MN ANALMGK YTHYKSRNLN FKFDMGKLSD DARISERESN SEALSKALSL TNCTTAMLKN LCFYSQESPQ SYNSVGPDTG RLK FSLSYK EQVGGNRELY IGDLRTKMFT RLIEDYFEAL SSQLSGSCLN NEKEFENAIL SMKLNVSMAH VSYSMDHSKW GPMM CPFLF LTVLQNLIFL SKDLQADIKG RDYLSTLLMW HMHKMVEIPF NVVSAMMKSF IKAQLGLRKK TKQSITEDFF YSNFQ IGVV PSHISSILDM GQGILHNTSD FYALITERFI NYAISCVCGG TIDAYTSSDD QISLFDQTLT ELLHRDPEEF RALMEF HYY MSDQLNKFVS PKSVIGRFVA EFKSRFFVWG DEVPLLTKFV AAALHNIKCK EPHQLAETID TIVDQSVANG VPVHLCN LI QIRTLSLLQY ARYPIDPFLL NCETDVRDWV DGNRSYRIMR QIEGLIPDAC SKIRSMLRRL YNRLKTGQLH EEFTTNYL S SEHLSSLKNL CELLGVEPPS ESDLEYSWLN LAAHHPLRMV LRQKIIYSGA VNLDDEKIPT IVKTIQNKLS STFTRGAQK LLSEAINKSA FQSSIASGFV GLCRTLGSKC VRGPNKENLY IKSIQSLITG TQGIELLTNS IGVQYWRVPL GLRNKSESVV SYFRPLLWD YMCISLSTAI ELGAWVLGDP KTTKALDFFK HNPCDYFPLK PTASKLLEDR VGLNHIIHSL RRLYPSVFEK H ILPFMSDL ASTKMKWSPR IKFLDLCVAL DVNCEALSLV SHIVKWKREE HYIVLSSELR FSHTRTHEPM VEERVVSTSD AV DNFMRQI YFESYVRPFV ATTRTLGSFT WFPHRTSIPE GEGLHRLGPF SSFVEKVIHK GVERPMFKHD LMMGYAWIDF DIE PARFNQ NQLIASGLVD SKFDSLEDFF DAVASLPTGS AKLSQTVRFR IKSQDASFRE SFAIHLDYIG SMNQQAKYLV HDVT AMYSG AVSPCVLSDC WRLVLSGPTF KGKPVWYVDT EVINEFLVDT NQLGHVTPVE VVVDMEKLQF AEYDFMLVGP CAEPV PLVV RRGGLWECEK KLASFTPVIQ DQDLEMFVRE VGDTSSDLLI RALSDMITDR LGLRMQWSGV DIVSTLRAAA PGNAEV LSA VLEVVDNWVE FKGYALCYSK SRGRVMVQSS SGKLRLKGRT CEELTEGGEH VEDIE

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Macromolecule #2: RING finger protein Z

MacromoleculeName: RING finger protein Z / type: protein_or_peptide / ID: 2 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Lassa mammarenavirus
Molecular weightTheoretical: 10.741461 KDa
Recombinant expressionOrganism: Escherichia coli (E. coli)
SequenceString:
MGNKQVKAPE ARNSPRASLI PDATHLGPQF CKSCWFENKG LVECNNHYLC LNCLTLLLGV SSRCPICKMP LPTRLRPSAA PTAPPAEAG DNTRPPPYSP

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Macromolecule #3: MANGANESE (II) ION

MacromoleculeName: MANGANESE (II) ION / type: ligand / ID: 3 / Number of copies: 1 / Formula: MN
Molecular weightTheoretical: 54.938 Da

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Macromolecule #4: ZINC ION

MacromoleculeName: ZINC ION / type: ligand / ID: 4 / Number of copies: 2 / Formula: ZN
Molecular weightTheoretical: 65.409 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

Concentration1 mg/mL
BufferpH: 7.5
GridModel: Quantifoil R1.2/1.3 / Material: COPPER / Mesh: 300 / Support film - Material: CARBON / Support film - topology: HOLEY / Pretreatment - Type: GLOW DISCHARGE / Pretreatment - Atmosphere: AIR
VitrificationCryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 277 K / Instrument: FEI VITROBOT MARK IV

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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 / Cs: 2.7 mm
Specialist opticsEnergy filter - Name: GIF Bioquantum / 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 K2 SUMMIT (4k x 4k) / Detector mode: SUPER-RESOLUTION / Digitization - Frames/image: 3-17 / Average electron dose: 60.0 e/Å2
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Particle selectionNumber selected: 1200000
CTF correctionSoftware - Name: CTFFIND (ver. 4.1)
Startup modelType of model: EMDB MAP
EMDB ID:

0706

Initial angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: RELION (ver. 3.0)
Final 3D classificationSoftware - Name: RELION (ver. 3.0)
Final angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: RELION (ver. 3.0)
Final reconstructionNumber classes used: 1 / Applied symmetry - Point group: C1 (asymmetric) / Algorithm: FOURIER SPACE / Resolution.type: BY AUTHOR / Resolution: 3.88 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: RELION (ver. 3.0) / Number images used: 273078
FSC plot (resolution estimation)

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Atomic model buiding 1

Initial model(PDB ID:

6klc

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5i72

)
RefinementSpace: REAL / Protocol: FLEXIBLE FIT / Target criteria: Correlation coefficient
Output model

PDB-7ckl:
Structure of Lassa virus polymerase bound to Z matrix protein

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