+Open data
-Basic information
Entry | Database: EMDB / ID: EMD-11118 | |||||||||
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Title | La Crosse virus polymerase at elongation mimicking stage | |||||||||
Map data | Cryo-EM map of La Crosse virus polymerase in elongation-mimicking state | |||||||||
Sample |
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Keywords | polymerase / transcription / replication / La Crosse virus | |||||||||
Function / homology | Function and homology information host cell endoplasmic reticulum / virion component / host cell endoplasmic reticulum-Golgi intermediate compartment / host cell Golgi apparatus / Hydrolases; Acting on ester bonds / hydrolase activity / RNA-directed RNA polymerase / viral RNA genome replication / RNA-dependent RNA polymerase activity / DNA-templated transcription ...host cell endoplasmic reticulum / virion component / host cell endoplasmic reticulum-Golgi intermediate compartment / host cell Golgi apparatus / Hydrolases; Acting on ester bonds / hydrolase activity / RNA-directed RNA polymerase / viral RNA genome replication / RNA-dependent RNA polymerase activity / DNA-templated transcription / nucleotide binding / RNA binding / metal ion binding Similarity search - Function | |||||||||
Biological species | La Crosse orthobunyavirus | |||||||||
Method | single particle reconstruction / cryo EM / Resolution: 3.02 Å | |||||||||
Authors | Arragain B / Effantin G | |||||||||
Funding support | France, 1 items
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Citation | Journal: Nat Commun / Year: 2020 Title: Pre-initiation and elongation structures of full-length La Crosse virus polymerase reveal functionally important conformational changes. Authors: Benoît Arragain / Grégory Effantin / Piotr Gerlach / Juan Reguera / Guy Schoehn / Stephen Cusack / Hélène Malet / Abstract: Bunyavirales is an order of segmented negative-strand RNA viruses comprising several life-threatening pathogens against which no effective treatment is currently available. Replication and ...Bunyavirales is an order of segmented negative-strand RNA viruses comprising several life-threatening pathogens against which no effective treatment is currently available. Replication and transcription of the RNA genome constitute essential processes performed by the virally encoded multi-domain RNA-dependent RNA polymerase. Here, we describe the complete high-resolution cryo-EM structure of La Crosse virus polymerase. It reveals the presence of key protruding C-terminal domains, notably the cap-binding domain, which undergoes large movements related to its role in transcription initiation, and a zinc-binding domain that displays a fold not previously observed. We capture the polymerase structure at pre-initiation and elongation states, uncovering the coordinated movement of the priming loop, mid-thumb ring linker and lid domain required for the establishment of a ten-base-pair template-product RNA duplex before strand separation into respective exit tunnels. These structural details and the observed dynamics of key functional elements will be instrumental for structure-based development of polymerase inhibitors. | |||||||||
History |
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-Structure visualization
Movie |
Movie viewer |
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Structure viewer | EM map: SurfViewMolmilJmol/JSmol |
Supplemental images |
-Downloads & links
-EMDB archive
Map data | emd_11118.map.gz | 4.5 MB | EMDB map data format | |
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Header (meta data) | emd-11118-v30.xml emd-11118.xml | 21.2 KB 21.2 KB | Display Display | EMDB header |
FSC (resolution estimation) | emd_11118_fsc.xml | 10.7 KB | Display | FSC data file |
Images | emd_11118.png | 177.3 KB | ||
Filedesc metadata | emd-11118.cif.gz | 8 KB | ||
Archive directory | http://ftp.pdbj.org/pub/emdb/structures/EMD-11118 ftp://ftp.pdbj.org/pub/emdb/structures/EMD-11118 | HTTPS FTP |
-Validation report
Summary document | emd_11118_validation.pdf.gz | 406.2 KB | Display | EMDB validaton report |
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Full document | emd_11118_full_validation.pdf.gz | 405.8 KB | Display | |
Data in XML | emd_11118_validation.xml.gz | 11.8 KB | Display | |
Data in CIF | emd_11118_validation.cif.gz | 15.4 KB | Display | |
Arichive directory | https://ftp.pdbj.org/pub/emdb/validation_reports/EMD-11118 ftp://ftp.pdbj.org/pub/emdb/validation_reports/EMD-11118 | HTTPS FTP |
-Related structure data
Related structure data | 6z8kMC 6z6bC 6z6gC C: citing same article (ref.) M: atomic model generated by this map |
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Similar structure data |
-Links
EMDB pages | EMDB (EBI/PDBe) / EMDataResource |
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-Map
File | Download / File: emd_11118.map.gz / Format: CCP4 / Size: 103 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES) | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
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Annotation | Cryo-EM map of La Crosse virus polymerase in elongation-mimicking state | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Voxel size | X=Y=Z: 0.826 Å | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Density |
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Symmetry | Space group: 1 | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Details | EMDB XML:
CCP4 map header:
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-Supplemental data
-Sample components
-Entire : La Crosse virus polymerase at elongation-mimicking stage
Entire | Name: La Crosse virus polymerase at elongation-mimicking stage |
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Components |
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-Supramolecule #1: La Crosse virus polymerase at elongation-mimicking stage
Supramolecule | Name: La Crosse virus polymerase at elongation-mimicking stage type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1-#4 |
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Molecular weight | Theoretical: 265 KDa |
-Supramolecule #2: La Crosse virus polymerase at elongation stage
Supramolecule | Name: La Crosse virus polymerase at elongation stage / type: complex / ID: 2 / Parent: 1 / Macromolecule list: #1-#3 |
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Source (natural) | Organism: La Crosse orthobunyavirus |
-Supramolecule #3: RNA-directed RNA polymerase L
Supramolecule | Name: RNA-directed RNA polymerase L / type: complex / ID: 3 / Parent: 1 / Macromolecule list: #4 |
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Source (natural) | Organism: La Crosse orthobunyavirus |
-Macromolecule #1: La Crosse virus 5' vRNA 1-10
Macromolecule | Name: La Crosse virus 5' vRNA 1-10 / type: rna / ID: 1 / Number of copies: 2 |
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Source (natural) | Organism: La Crosse orthobunyavirus |
Molecular weight | Theoretical: 3.217956 KDa |
Sequence | String: AGUAGUGUGC |
-Macromolecule #2: La Crosse virus 5' vRNA (9-16)
Macromolecule | Name: La Crosse virus 5' vRNA (9-16) / type: rna / ID: 2 / Number of copies: 1 |
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Source (natural) | Organism: La Crosse orthobunyavirus |
Molecular weight | Theoretical: 2.509577 KDa |
Sequence | String: GCUACCAA |
-Macromolecule #3: La Crosse virus 3' vRNA (1-16)
Macromolecule | Name: La Crosse virus 3' vRNA (1-16) / type: rna / ID: 3 / Number of copies: 2 |
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Source (natural) | Organism: La Crosse orthobunyavirus |
Molecular weight | Theoretical: 5.060023 KDa |
Sequence | String: UUGGUAGUAC ACUACU |
-Macromolecule #4: RNA-directed RNA polymerase L
Macromolecule | Name: RNA-directed RNA polymerase L / type: protein_or_peptide / ID: 4 / Number of copies: 1 / Enantiomer: LEVO / EC number: RNA-directed RNA polymerase |
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Source (natural) | Organism: La Crosse orthobunyavirus |
Molecular weight | Theoretical: 266.047469 KDa |
Recombinant expression | Organism: Trichoplusia ni (cabbage looper) |
Sequence | String: MGHHHHHHDY DIPTTENLYF QGMDYQEYQQ FLARINTARD ACVAKDIDVD LLMARHDYFG RELCKSLNIE YRNDVPFIDI ILDIRPEVD PLTIDAPHIT PDNYLYINNV LYIIDYKVSV SNESSVITYD KYYELTRDIS DRLSIPIEIV IIRIDPVSRD L HINSDRFK ...String: MGHHHHHHDY DIPTTENLYF QGMDYQEYQQ FLARINTARD ACVAKDIDVD LLMARHDYFG RELCKSLNIE YRNDVPFIDI ILDIRPEVD PLTIDAPHIT PDNYLYINNV LYIIDYKVSV SNESSVITYD KYYELTRDIS DRLSIPIEIV IIRIDPVSRD L HINSDRFK ELYPTIVVDI NFNQFFDLKQ LLYEKFGDDE EFLLKVAHGD FTLTAPWCKT GCPEFWKHPI YKEFKMSMPV PE RRLFEES VKFNAYESER WNTNLVKIRE YTKKDYSEHI SKSAKNIFLA SGFYKQPNKN EISEGWTLMV ERVQDQREIS KSL HDQKPS IHFIWGAHNP GNSNNATFKL ILLSKSLQSI KGISTYTEAF KSLGKMMDIG DKAIEYEEFC MSLKSKARSS WKQI MNKKL EPKQINNALV LWEQQFMINN DLIDKSEKLK LFKNFCGIGK HKQFKNKMLE DLEVSKPKIL DFDDANMYLA SLTMM EQSK KILSKSNGLK PDNFILNEFG SRIKDANKET YDNMHKIFET GYWQCISDFS TLMKNILSVS QYNRHNTFRI AMCANN NVF AIVFPSADIK TKKATVVYSI IVLHKEEENI FNPGCLHGTF KCMNGYISIS RAIRLDKERC QRIVSSPGLF LTTCLLF KH DNPTLVMSDI MNFSIYTSLS ITKSVLSLTE PARYMIMNSL AISSNVKDYI AEKFSPYTKT LFSVYMTRLI KNACFDAY D QRQRVQLRDI YLSDYDITQK GIKDNRELTS IWFPGSVTLK EYLTQIYLPF YFNAKGLHEK HHVMVDLAKT ILEIECEQR ENIKEIWSTN CTKQTVNLKI LIHSLCKNLL ADTSRHNHLR NRIENRNNFR RSITTISTFT SSKSCLKIGD FRKEKELQSV KQKKILEVQ SRKMRLANPM FVTDEQVCLE VGHCNYEMLR NAMPNYTDYI STKVFDRLYE LLDKKVLTDK PVIEQIMDMM I DHKKFYFT FFNKGQKTSK DREIFVGEYE AKMCMYAVER IAKERCKLNP DEMISEPGDG KLKVLEQKSE QEIRFLVETT RQ KNREIDE AIEALATEGY ESNLGKIEKL SLGKAKGLKM EINADMSKWS AQDVFYKYFW LIALDPILYP QEKERILYFM CNY MDKELI LPDELLFNLL DQKVAYQNDI IATMTNQLNS NTVLIKRNWL QGNFNYTSSY VHSCAMSVYK EILKEAITLL DGSI LVNSL VHSDDNQTSI TIVQDKMEND KIIDFAMKEF ERACLTFGCQ ANMKKTYVTN CIKEFVSLFN LYGEPFSIYG RFLLT SVGD CAYIGPYEDL ASRISSAQTA IKHGCPPSLA WVSIAISHWM TSLTYNMLPG QSNDPIDYFP AENRKDIPIE LNGVLD APL SMISTVGLES GNLYFLIKLL SKYTPVMQKR ESVVNQIAEV KNWKVEDLTD NEIFRLKILR YLVLDAEMDP SDIMGET SD MRGRSILTPR KFTTAGSLRK LYSFSKYQDR LSSPGGMVEL FTYLLEKPEL LVTKGEDMKD YMESVIFRYN SKRFKESL S IQNPAQLFIE QILFSHKPVI DFSGIRDKYI NLHDSRALEK EPDILGKVTF TEAYRLLMRD LSSLELTNDD IQVIYSYII LNDPMMITIA NTHILSIYGS PQRRMGMSCS TMPEFRNLKL IHHSPALVLR AYSKNNPDIQ GADPTEMARD LVHLKEFVEN TNLEEKMKV RIAMNEAEKG QRDIVFELKE MTRFYQVCYE YVKSTEHKIK VFILPAKSYT TTDFCSLMQG NLIKDKEWYT V HYLKQILS GGHKAIMQHN ATSEQNIAFE CFKLITHFAD SFIDSLSRSA FLQLIIDEFS YKDVKVSKLY DIIKNGYNRT DF IPLLFRT GDLRQADLDK YDAMKSHERV TWNDWQTSRH LDMGSINLTI TGYNRSITII GEDNKLTYAE LCLTRKTPEN ITI SGRKLL GSRHGLKFEN MSKIQTYPGN YYITYRKKDR HQFVYQIHSH ESITRRNEEH MAIRTRIYNE ITPVCVVNVA EVDG DQRIL IRSLDYLNND IFSLSRIKVG LDEFATIKKA HFSKMVSFEG PPIKTGLLDL TELMKSQDLL NLNYDNIRNS NLISF SKLI CCEGSDNIND GLEFLSDDPM NFTEGEAIHS TPIFNIYYSK RGERHMTYRN AIKLLIERET KIFEEAFTFS ENGFIS PEN LGCLEAVVSL IKLLKTNEWS TVIDKCIHIC LIKNGMDHMY HSFDVPKCFM GNPITRDINW VMFREFINSL PGTDIPP WN VMTENFKKKC IALINSKFET QRDFSEFTKL MKKEGGRSNI EFD UniProtKB: RNA-directed RNA polymerase L |
-Macromolecule #5: ZINC ION
Macromolecule | Name: ZINC ION / type: ligand / ID: 5 / Number of copies: 1 / Formula: ZN |
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Molecular weight | Theoretical: 65.409 Da |
-Macromolecule #6: MAGNESIUM ION
Macromolecule | Name: MAGNESIUM ION / type: ligand / ID: 6 / Number of copies: 1 / Formula: MG |
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Molecular weight | Theoretical: 24.305 Da |
-Experimental details
-Structure determination
Method | cryo EM |
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Processing | single particle reconstruction |
Aggregation state | particle |
-Sample preparation
Concentration | 0.2 mg/mL | ||||||||
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Buffer | pH: 8 Component:
Details: 20 mM Tris-HCl pH 8, 150 mM NaCl, 2 mM TCEP | ||||||||
Grid | Model: UltrAuFoil / Material: GOLD / Mesh: 300 / Support film - Material: GOLD / Support film - topology: HOLEY / Pretreatment - Type: GLOW DISCHARGE / Pretreatment - Time: 60 sec. / Details: 30mA | ||||||||
Vitrification | Cryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 293 K / Instrument: FEI VITROBOT MARK IV / Details: blot time: 2 sec, blot force: 1. |
-Electron microscopy
Microscope | FEI TITAN KRIOS |
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Temperature | Min: 90.0 K / Max: 90.0 K |
Specialist optics | Energy filter - Name: GIF Quantum LS / Energy filter - Slit width: 20 eV |
Image recording | Film or detector model: GATAN K2 SUMMIT (4k x 4k) / Detector mode: SUPER-RESOLUTION / Number grids imaged: 2 / Number real images: 16498 / Average electron dose: 1.25 e/Å2 |
Electron beam | Acceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN |
Electron optics | Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Cs: 2.7 mm / Nominal defocus max: -3.5 µm / Nominal defocus min: -0.8 µm / Nominal magnification: 165000 |
Sample stage | Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Cooling holder cryogen: NITROGEN |
Experimental equipment | Model: Titan Krios / Image courtesy: FEI Company |