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- PDB-8xpo: Cryo-EM structure of Lassa virus RdRP elongation complex with the... -

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Basic information

Entry
Database: PDB / ID: 8xpo
TitleCryo-EM structure of Lassa virus RdRP elongation complex with the NTP form of compound HNC-1664 bound in the active site
Components
  • RNA (14-MER)
  • RNA (32-MER)
  • RNA (9-MER)
  • RNA-directed RNA polymerase L
KeywordsREPLICATION / Lassa virus / RNA-dependent RNA polymerase / elongation complex / nucleoside analog
Function / homology
Function and homology information


negative stranded viral RNA replication / cap snatching / virion component / host cell / Hydrolases; Acting on ester bonds / host cell cytoplasm / hydrolase activity / RNA-directed RNA polymerase / nucleotide binding / RNA-directed RNA polymerase activity / metal ion binding
Similarity search - Function
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
: / RNA / RNA (> 10) / RNA-directed RNA polymerase L
Similarity search - Component
Biological speciesLassa virus Josiah
synthetic construct (others)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 3.02 Å
AuthorsJing, X. / Gong, P.
Funding support China, 2items
OrganizationGrant numberCountry
National Natural Science Foundation of China (NSFC)(2022YFC2303300) China
National Natural Science Foundation of China (NSFC)U23A20147 China
CitationJournal: Nat Commun / Year: 2024
Title: An adenosine analog shows high antiviral potency against coronavirus and arenavirus mainly through an unusual base pairing mode.
Authors: Xiaoying Jia / Xuping Jing / Ming Li / Minli Gao / Yao Zhong / Entao Li / Yang Liu / Rui Li / Guoqiang Yao / Qiaojie Liu / Minmin Zhou / Yuxia Hou / Linfeng An / Yibao Hong / Shanshan Li / ...Authors: Xiaoying Jia / Xuping Jing / Ming Li / Minli Gao / Yao Zhong / Entao Li / Yang Liu / Rui Li / Guoqiang Yao / Qiaojie Liu / Minmin Zhou / Yuxia Hou / Linfeng An / Yibao Hong / Shanshan Li / Jiancun Zhang / Wei Wang / Kaiming Zhang / Peng Gong / Sandra Chiu /
Abstract: By targeting the essential viral RNA-dependent RNA polymerase (RdRP), nucleoside analogs (NAs) have exhibited great potential in antiviral therapy for RNA virus-related diseases. However, most ribose- ...By targeting the essential viral RNA-dependent RNA polymerase (RdRP), nucleoside analogs (NAs) have exhibited great potential in antiviral therapy for RNA virus-related diseases. However, most ribose-modified NAs do not present broad-spectrum features, likely due to differences in ribose-RdRP interactions across virus families. Here, we show that HNC-1664, an adenosine analog with modifications both in ribose and base, has broad-spectrum antiviral activity against positive-strand coronaviruses and negative-strand arenaviruses. Importantly, treatment with HNC-1664 demonstrate anti-SARS-CoV-2 efficacy in infected K18-human ACE2 mice, with reduced viral titer and mortality, as well as improved lung injury. Enzymology data demonstrate that HNC-1664 inhibits RNA synthesis mainly at the pre-catalysis stage. The cryo-EM structures of HNC-1664-bound RdRP-RNA complexes from both SARS-CoV-2 and LASV reveal an unusual base pairing mode of HNC-1664 in part due to its base modification, thus revealing its great potency in binding but not catalysis. Under certain circumstances, 1664-TP can be slowly incorporated by RdRP through regular Watson-Crick base pairing, as evidenced by enzymology data and an HNC-1664-incorporated crystal structure of the RdRP-RNA complex. Overall, HNC-1664 achieves broad-spectrum characteristics by favoring an alternative base pairing strategy to non-catalytically block RNA synthesis, providing a novel concept for the rational development of NA drugs.
History
DepositionJan 4, 2024Deposition site: PDBJ / Processing site: PDBJ
Revision 1.0Dec 4, 2024Provider: repository / Type: Initial release
Revision 1.1Jan 15, 2025Group: Data collection / Database references / Category: citation / citation_author / em_admin
Item: _citation.journal_volume / _citation.page_first ..._citation.journal_volume / _citation.page_first / _citation.page_last / _citation.pdbx_database_id_DOI / _citation.pdbx_database_id_PubMed / _citation.title / _citation_author.identifier_ORCID / _citation_author.name / _em_admin.last_update

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

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Assembly

Deposited unit
L: RNA-directed RNA polymerase L
A: RNA (14-MER)
T: RNA (32-MER)
P: RNA (9-MER)
hetero molecules


Theoretical massNumber of molelcules
Total (without water)273,5335
Polymers272,9004
Non-polymers6331
Water00
1


  • Idetical with deposited unit
  • defined by author
  • Evidence: electron microscopy
TypeNameSymmetry operationNumber
identity operation1_5551

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Components

#1: Protein RNA-directed RNA polymerase L / Protein L / Large structural protein / Replicase / Transcriptase


Mass: 255200.422 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Lassa virus Josiah / Gene: L / Production host: Pichia (fungus)
References: UniProt: Q6Y630, RNA-directed RNA polymerase, Hydrolases; Acting on ester bonds
#2: RNA chain RNA (14-MER)


Mass: 4516.763 Da / Num. of mol.: 1 / Source method: obtained synthetically / Source: (synth.) synthetic construct (others)
#3: RNA chain RNA (32-MER)


Mass: 10374.283 Da / Num. of mol.: 1 / Source method: obtained synthetically / Source: (synth.) synthetic construct (others)
#4: RNA chain RNA (9-MER)


Mass: 2808.703 Da / Num. of mol.: 1 / Source method: obtained synthetically / Source: (synth.) synthetic construct (others)
#5: Chemical ChemComp-A1LVZ / [[(2~{R},3~{R},4~{S},5~{R})-4-fluoranyl-5-(5-iodanyl-4-methyl-pyrrolo[2,3-d]pyrimidin-7-yl)-3-oxidanyl-oxolan-2-yl]methoxy-oxidanyl-phosphoryl] phosphono hydrogen phosphate


Mass: 633.092 Da / Num. of mol.: 1 / Source method: obtained synthetically / Formula: C12H16FIN3O12P3 / Feature type: SUBJECT OF INVESTIGATION
Has ligand of interestY
Has protein modificationN

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Experimental details

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Experiment

ExperimentMethod: ELECTRON MICROSCOPY
EM experimentAggregation state: PARTICLE / 3D reconstruction method: single particle reconstruction

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Sample preparation

Component
IDNameTypeEntity IDParent-IDSource
1Lassa virus RdRP elongation complex with the NTP form of compound 13b bound in the active siteCOMPLEX#1-#40MULTIPLE SOURCES
2Lassa virus L proteinCOMPLEX#11RECOMBINANT
3RNACOMPLEX#2-#41SYNTHETIC
Molecular weightValue: 0.27 MDa / Experimental value: NO
Source (natural)
IDEntity assembly-IDOrganismNcbi tax-ID
22Lassa virus Josiah11622
33synthetic construct (others)32630
Source (recombinant)Organism: Pichia (fungus)
Buffer solutionpH: 7
SpecimenEmbedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
Specimen supportGrid material: COPPER / Grid mesh size: 300 divisions/in. / Grid type: Quantifoil R1.2/1.3
VitrificationInstrument: FEI VITROBOT MARK IV / Cryogen name: ETHANE

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Electron microscopy imaging

MicroscopyModel: JEOL CRYO ARM 300
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Illumination mode: FLOOD BEAM
Electron lensMode: BRIGHT FIELD / Nominal defocus max: 2500 nm / Nominal defocus min: 500 nm
Image recordingElectron dose: 40 e/Å2 / Detector mode: SUPER-RESOLUTION / Film or detector model: GATAN K3 (6k x 4k)

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Processing

EM softwareName: PHENIX / Version: 1.18.2_3874: / Category: model refinement
CTF correctionType: PHASE FLIPPING ONLY
3D reconstructionResolution: 3.02 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 225955 / Symmetry type: POINT
Atomic model buildingProtocol: AB INITIO MODEL
Refine LS restraints
Refine-IDTypeDev idealNumber
ELECTRON MICROSCOPYf_bond_d0.01314490
ELECTRON MICROSCOPYf_angle_d0.84419791
ELECTRON MICROSCOPYf_dihedral_angle_d11.482259
ELECTRON MICROSCOPYf_chiral_restr0.0532314
ELECTRON MICROSCOPYf_plane_restr0.0052370

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