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- EMDB-32599: Cryo-EM structure of tetrameric TLR3 in complex with dsRNA (90 bp) -

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Basic information

Entry
Database: EMDB / ID: EMD-32599
TitleCryo-EM structure of tetrameric TLR3 in complex with dsRNA (90 bp)
Map data
Sample
  • Complex: Tetrameric TLR3 in complex with dsRNA (90 bp)
    • Protein or peptide: Toll-like receptor 3
    • RNA: RNA (81-MER)
    • RNA: RNA (81-MER)
  • Ligand: 2-acetamido-2-deoxy-beta-D-glucopyranose
Keywordsinnate immunity / IMMUNE SYSTEM / IMMUNE SYSTEM-RNA complex
Function / homology
Function and homology information


type III interferon production / positive regulation of type III interferon production / response to dsRNA / regulation of dendritic cell cytokine production / inflammatory response to wounding / toll-like receptor 3 signaling pathway / necroptotic signaling pathway / positive regulation of cytokine production involved in inflammatory response / positive regulation of macrophage cytokine production / pattern recognition receptor activity ...type III interferon production / positive regulation of type III interferon production / response to dsRNA / regulation of dendritic cell cytokine production / inflammatory response to wounding / toll-like receptor 3 signaling pathway / necroptotic signaling pathway / positive regulation of cytokine production involved in inflammatory response / positive regulation of macrophage cytokine production / pattern recognition receptor activity / toll-like receptor signaling pathway / response to exogenous dsRNA / cellular response to exogenous dsRNA / positive regulation of interferon-alpha production / positive regulation of type I interferon production / cellular response to interferon-beta / positive regulation of chemokine production / extrinsic apoptotic signaling pathway / JNK cascade / positive regulation of interleukin-12 production / positive regulation of interferon-beta production / positive regulation of interleukin-8 production / microglial cell activation / positive regulation of JNK cascade / response to virus / defense response / cellular response to virus / cellular response to type II interferon / positive regulation of interleukin-6 production / positive regulation of non-canonical NF-kappaB signal transduction / cellular response to mechanical stimulus / male gonad development / positive regulation of angiogenesis / MAPK cascade / transmembrane signaling receptor activity / positive regulation of type II interferon production / cellular response to xenobiotic stimulus / double-stranded RNA binding / positive regulation of tumor necrosis factor production / signaling receptor activity / defense response to virus / positive regulation of canonical NF-kappaB signal transduction / early endosome / endosome membrane / inflammatory response / positive regulation of apoptotic process / innate immune response / endoplasmic reticulum membrane / positive regulation of gene expression / cell surface / positive regulation of transcription by RNA polymerase II / identical protein binding / plasma membrane / cytoplasm
Similarity search - Function
Toll-like receptor 3 trans-membrane domain / Toll-like receptor 3 trans-membrane domain / Toll-like receptor / TIR domain / Leucine Rich Repeat / Cysteine-rich flanking region, C-terminal / Leucine rich repeat C-terminal domain / Toll - interleukin 1 - resistance / Leucine-rich repeat, SDS22-like subfamily / TIR domain profile. ...Toll-like receptor 3 trans-membrane domain / Toll-like receptor 3 trans-membrane domain / Toll-like receptor / TIR domain / Leucine Rich Repeat / Cysteine-rich flanking region, C-terminal / Leucine rich repeat C-terminal domain / Toll - interleukin 1 - resistance / Leucine-rich repeat, SDS22-like subfamily / TIR domain profile. / Toll/interleukin-1 receptor homology (TIR) domain / Toll/interleukin-1 receptor homology (TIR) domain superfamily / Leucine rich repeat / Leucine-rich repeat, typical subtype / Leucine-rich repeats, typical (most populated) subfamily / Leucine-rich repeat profile. / Leucine-rich repeat / Leucine-rich repeat domain superfamily
Similarity search - Domain/homology
Toll-like receptor 3
Similarity search - Component
Biological speciesMus musculus (house mouse) / synthetic construct (others)
Methodsingle particle reconstruction / cryo EM / Resolution: 3.2 Å
AuthorsSakaniwa K / Ohto U
Funding support Japan, 3 items
OrganizationGrant numberCountry
Ministry of Education, Culture, Sports, Science and Technology (Japan)19J21830 Japan
Ministry of Education, Culture, Sports, Science and Technology (Japan)19H03164 Japan
Ministry of Education, Culture, Sports, Science and Technology (Japan)19H00976 Japan
CitationJournal: Nat Commun / Year: 2023
Title: TLR3 forms a laterally aligned multimeric complex along double-stranded RNA for efficient signal transduction.
Authors: Kentaro Sakaniwa / Akiko Fujimura / Takuma Shibata / Hideki Shigematsu / Toru Ekimoto / Masaki Yamamoto / Mitsunori Ikeguchi / Kensuke Miyake / Umeharu Ohto / Toshiyuki Shimizu /
Abstract: Toll-like receptor 3 (TLR3) is a member of the TLR family, which plays an important role in the innate immune system and is responsible for recognizing viral double-stranded RNA (dsRNA). Previous ...Toll-like receptor 3 (TLR3) is a member of the TLR family, which plays an important role in the innate immune system and is responsible for recognizing viral double-stranded RNA (dsRNA). Previous biochemical and structural studies have revealed that a minimum length of approximately 40-50 base pairs of dsRNA is necessary for TLR3 binding and dimerization. However, efficient TLR3 activation requires longer dsRNA and the molecular mechanism underlying its dsRNA length-dependent activation remains unknown. Here, we report cryo-electron microscopy analyses of TLR3 complexed with longer dsRNA. TLR3 dimers laterally form a higher multimeric complex along dsRNA, providing the basis for cooperative binding and efficient signal transduction.
History
DepositionJan 14, 2022-
Header (metadata) releaseJan 25, 2023-
Map releaseJan 25, 2023-
UpdateAug 30, 2023-
Current statusAug 30, 2023Processing site: PDBj / Status: Released

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Structure visualization

Supplemental images

emd_32599.png

Downloads & links

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Map

FileDownload / File: emd_32599.map.gz / Format: CCP4 / Size: 83.7 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Voxel sizeX=Y=Z: 1.128 Å
Density
Contour LevelBy AUTHOR: 0.05
Minimum - Maximum-0.17174713 - 0.4463007
Average (Standard dev.)0.00074188976 (±0.017022053)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions280280280
Spacing280280280
CellA=B=C: 315.84 Å
α=β=γ: 90.0 °

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Supplemental data

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Sample components

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Entire : Tetrameric TLR3 in complex with dsRNA (90 bp)

EntireName: Tetrameric TLR3 in complex with dsRNA (90 bp)
Components
  • Complex: Tetrameric TLR3 in complex with dsRNA (90 bp)
    • Protein or peptide: Toll-like receptor 3
    • RNA: RNA (81-MER)
    • RNA: RNA (81-MER)
  • Ligand: 2-acetamido-2-deoxy-beta-D-glucopyranose

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Supramolecule #1: Tetrameric TLR3 in complex with dsRNA (90 bp)

SupramoleculeName: Tetrameric TLR3 in complex with dsRNA (90 bp) / type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1-#3
Source (natural)Organism: Mus musculus (house mouse)

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Macromolecule #1: Toll-like receptor 3

MacromoleculeName: Toll-like receptor 3 / type: protein_or_peptide / ID: 1 / Number of copies: 4 / Enantiomer: LEVO
Source (natural)Organism: Mus musculus (house mouse)
Molecular weightTheoretical: 77.365852 KDa
Recombinant expressionOrganism: Homo sapiens (human)
SequenceString: TNQCTVRYNV ADCSHLKLTH IPDDLPSNIT VLNLTHNQLR RLPPTNFTRY SQLAILDAGF NSISKLEPEL CQILPLLKVL NLQHNELSQ ISDQTFVFCT NLTELDLMSN SIHKIKSNPF KNQKNLIKLD LSHNGLSSTK LGTGVQLENL QELLLAKNKI L ALRSEELE ...String:
TNQCTVRYNV ADCSHLKLTH IPDDLPSNIT VLNLTHNQLR RLPPTNFTRY SQLAILDAGF NSISKLEPEL CQILPLLKVL NLQHNELSQ ISDQTFVFCT NLTELDLMSN SIHKIKSNPF KNQKNLIKLD LSHNGLSSTK LGTGVQLENL QELLLAKNKI L ALRSEELE FLGNSSLRKL DLSSNPLKEF SPGCFQTIGK LFALLLNNAQ LNPHLTEKLC WELSNTSIQN LSLANNQLLA TS ESTFSGL KWTNLTQLDL SYNNLHDVGN GSFSYLPSLR YLSLEYNNIQ RLSPRSFYGL SNLRYLSLKR AFTKQSVSLA SHP NIDDFS FQWLKYLEYL NMDDNNIPST KSNTFTGLVS LKYLSLSKTF TSLQTLTNET FVSLAHSPLL TLNLTKNHIS KIAN GTFSW LGQLRILDLG LNEIEQKLSG QEWRGLRNIF EIYLSYNKYL QLSTSSFALV PSLQRLMLRR VALKNVDISP SPFRP LRNL TILDLSNNNI ANINEDLLEG LENLEILDFQ HNNLARLWKR ANPGGPVNFL KGLSHLHILN LESNGLDEIP VGVFKN LFE LKSINLGLNN LNKLEPFIFD DQTSLRSLNL QKNLITSVEK DVFGPPFQNL NSLDMRFNPF DCTCESISWF VNWINQT HT NISELSTHYL CNTPHHYYGF PLKLFDTSSC KDSAPFEL

UniProtKB: Toll-like receptor 3

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Macromolecule #2: RNA (81-MER)

MacromoleculeName: RNA (81-MER) / type: rna / ID: 2 / Number of copies: 1
Source (natural)Organism: synthetic construct (others)
Molecular weightTheoretical: 25.699127 KDa
SequenceString:
AAAAAAAAAA AAAAAAAAAA AAAAAAAAAA AAAAAAAAAA AUUUUUUUUU UUUUUUUUUU UUUUUUUUUU UUUUUUUUUU U

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Macromolecule #3: RNA (81-MER)

MacromoleculeName: RNA (81-MER) / type: rna / ID: 3 / Number of copies: 1
Source (natural)Organism: synthetic construct (others)
Molecular weightTheoretical: 25.676088 KDa
SequenceString:
AAAAAAAAAA AAAAAAAAAA AAAAAAAAAA AAAAAAAAAA UUUUUUUUUU UUUUUUUUUU UUUUUUUUUU UUUUUUUUUU U

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Macromolecule #6: 2-acetamido-2-deoxy-beta-D-glucopyranose

MacromoleculeName: 2-acetamido-2-deoxy-beta-D-glucopyranose / type: ligand / ID: 6 / Number of copies: 12 / Formula: NAG
Molecular weightTheoretical: 221.208 Da
Chemical component information

ChemComp-NAG:
2-acetamido-2-deoxy-beta-D-glucopyranose / N-Acetylglucosamine

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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

Concentration0.2 mg/mL
BufferpH: 5.3 / Details: 25 mM MES-NaOH pH 5.3, 0.3 M NaCl
Sugar embeddingMaterial: ice
VitrificationCryogen name: ETHANE / Instrument: FEI VITROBOT MARK IV

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Electron microscopy

MicroscopeJEOL CRYO ARM 300
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELDBright-field microscopy / Nominal defocus max: 2.4 µm / Nominal defocus min: 1.4000000000000001 µm
Image recordingFilm or detector model: GATAN K3 (6k x 4k) / Average electron dose: 50.0 e/Å2

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Image processing

Startup modelType of model: PDB ENTRY
PDB model - PDB ID:

3ciy

Initial angle assignmentType: MAXIMUM LIKELIHOOD
Final angle assignmentType: MAXIMUM LIKELIHOOD
Final reconstructionResolution.type: BY AUTHOR / Resolution: 3.2 Å / Resolution method: FSC 0.143 CUT-OFF / Number images used: 274002

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