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- EMDB-9947: Structure of ASC CARD filament -

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Basic information

Entry
Database: EMDB / ID: EMD-9947
TitleStructure of ASC CARD filament
Map dataNone
Sample
  • Complex: ASC CARD filament
    • Protein or peptide: Apoptosis-associated speck-like protein containing a CARD
Keywordsfilament / SIGNALING PROTEIN
Function / homology
Function and homology information


NLRP6 inflammasome complex / myosin I binding / Pyrin domain binding / myeloid dendritic cell activation involved in immune response / positive regulation of antigen processing and presentation of peptide antigen via MHC class II / negative regulation of protein serine/threonine kinase activity / regulation of intrinsic apoptotic signaling pathway / myeloid dendritic cell activation / IkappaB kinase complex / The AIM2 inflammasome ...NLRP6 inflammasome complex / myosin I binding / Pyrin domain binding / myeloid dendritic cell activation involved in immune response / positive regulation of antigen processing and presentation of peptide antigen via MHC class II / negative regulation of protein serine/threonine kinase activity / regulation of intrinsic apoptotic signaling pathway / myeloid dendritic cell activation / IkappaB kinase complex / The AIM2 inflammasome / AIM2 inflammasome complex / icosanoid biosynthetic process / NLRP1 inflammasome complex / macropinocytosis / canonical inflammasome complex / interleukin-6 receptor binding / NLRP3 inflammasome complex assembly / BMP receptor binding / positive regulation of adaptive immune response / NLRP3 inflammasome complex / cysteine-type endopeptidase activator activity / CLEC7A/inflammasome pathway / osmosensory signaling pathway / negative regulation of interferon-beta production / regulation of tumor necrosis factor-mediated signaling pathway / positive regulation of extrinsic apoptotic signaling pathway / pattern recognition receptor signaling pathway / positive regulation of macrophage cytokine production / negative regulation of NF-kappaB transcription factor activity / pattern recognition receptor activity / tropomyosin binding / pyroptotic inflammatory response / positive regulation of actin filament polymerization / positive regulation of release of cytochrome c from mitochondria / positive regulation of activated T cell proliferation / intrinsic apoptotic signaling pathway by p53 class mediator / The NLRP3 inflammasome / positive regulation of interleukin-10 production / cellular response to interleukin-1 / intrinsic apoptotic signaling pathway in response to DNA damage by p53 class mediator / positive regulation of T cell migration / positive regulation of DNA-binding transcription factor activity / Purinergic signaling in leishmaniasis infection / positive regulation of chemokine production / positive regulation of defense response to virus by host / negative regulation of canonical NF-kappaB signal transduction / negative regulation of cytokine production involved in inflammatory response / activation of innate immune response / intrinsic apoptotic signaling pathway / positive regulation of phagocytosis / positive regulation of interleukin-1 beta production / tumor necrosis factor-mediated signaling pathway / positive regulation of interleukin-8 production / positive regulation of JNK cascade / apoptotic signaling pathway / positive regulation of non-canonical NF-kappaB signal transduction / positive regulation of NF-kappaB transcription factor activity / protein homooligomerization / positive regulation of T cell activation / regulation of protein stability / positive regulation of interleukin-6 production / positive regulation of type II interferon production / positive regulation of inflammatory response / positive regulation of tumor necrosis factor production / SARS-CoV-1 activates/modulates innate immune responses / cellular response to tumor necrosis factor / azurophil granule lumen / cellular response to lipopolysaccharide / regulation of inflammatory response / protease binding / secretory granule lumen / defense response to Gram-negative bacterium / defense response to virus / microtubule / transmembrane transporter binding / positive regulation of ERK1 and ERK2 cascade / positive regulation of canonical NF-kappaB signal transduction / protein dimerization activity / defense response to Gram-positive bacterium / regulation of autophagy / positive regulation of apoptotic process / inflammatory response / Golgi membrane / innate immune response / neuronal cell body / apoptotic process / Neutrophil degranulation / nucleolus / enzyme binding / endoplasmic reticulum / signal transduction / protein homodimerization activity / protein-containing complex / mitochondrion / extracellular region / nucleoplasm / identical protein binding / nucleus / cytoplasm / cytosol
Similarity search - Function
CARD8/ASC/NALP1, CARD domain / : / DAPIN domain / DAPIN domain profile. / PAAD/DAPIN/Pyrin domain / PAAD/DAPIN/Pyrin domain / CARD domain / CARD caspase recruitment domain profile. / Caspase recruitment domain / Death-like domain superfamily
Similarity search - Domain/homology
Apoptosis-associated speck-like protein containing a CARD
Similarity search - Component
Biological speciesHomo sapiens (human)
Methodhelical reconstruction / cryo EM / Resolution: 4.1 Å
AuthorsGong Q / Xu C
Funding support Singapore, 1 items
OrganizationGrant numberCountry
Ministry of Education (Singapore)tier2 and tier1 Singapore
CitationJournal: Nat Commun / Year: 2021
Title: Structural basis for distinct inflammasome complex assembly by human NLRP1 and CARD8.
Authors: Qin Gong / Kim Robinson / Chenrui Xu / Phuong Thao Huynh / Kelvin Han Chung Chong / Eddie Yong Jun Tan / Jiawen Zhang / Zhao Zhi Boo / Daniel Eng Thiam Teo / Kenneth Lay / Yaming Zhang / ...Authors: Qin Gong / Kim Robinson / Chenrui Xu / Phuong Thao Huynh / Kelvin Han Chung Chong / Eddie Yong Jun Tan / Jiawen Zhang / Zhao Zhi Boo / Daniel Eng Thiam Teo / Kenneth Lay / Yaming Zhang / John Soon Yew Lim / Wah Ing Goh / Graham Wright / Franklin L Zhong / Bruno Reversade / Bin Wu /
Abstract: Nod-like receptor (NLR) proteins activate pyroptotic cell death and IL-1 driven inflammation by assembling and activating the inflammasome complex. Closely related sensor proteins NLRP1 and CARD8 ...Nod-like receptor (NLR) proteins activate pyroptotic cell death and IL-1 driven inflammation by assembling and activating the inflammasome complex. Closely related sensor proteins NLRP1 and CARD8 undergo unique auto-proteolysis-dependent activation and are implicated in auto-inflammatory diseases; however, their mechanisms of activation are not understood. Here we report the structural basis of how the activating domains (FIIND-CARD) of NLRP1 and CARD8 self-oligomerize to assemble distinct inflammasome complexes. Recombinant FIIND-CARD of NLRP1 forms a two-layered filament, with an inner core of oligomerized CARD surrounded by an outer ring of FIIND. Biochemically, self-assembled NLRP1-CARD filaments are sufficient to drive ASC speck formation in cultured human cells-a process that is greatly enhanced by NLRP1-FIIND which forms oligomers in vitro. The cryo-EM structures of NLRP1-CARD and CARD8-CARD filaments, solved here at 3.7 Å, uncover unique structural features that enable NLRP1 and CARD8 to discriminate between ASC and pro-caspase-1. In summary, our findings provide structural insight into the mechanisms of activation for human NLRP1 and CARD8 and reveal how highly specific signaling can be achieved by heterotypic CARD interactions within the inflammasome complexes.
History
DepositionJun 14, 2019-
Header (metadata) releaseSep 16, 2020-
Map releaseSep 16, 2020-
UpdateMar 27, 2024-
Current statusMar 27, 2024Processing site: PDBj / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 0.0135
  • Imaged by UCSF Chimera
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  • Surface view colored by cylindrical radius
  • Surface level: 0.0135
  • Imaged by UCSF Chimera
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  • Surface view with fitted model
  • Atomic models: PDB-6k99
  • Surface level: 0.0135
  • Imaged by UCSF Chimera
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  • Simplified surface model + fitted atomic model
  • Atomic modelsPDB-6k99
  • Imaged by Jmol
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Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

emd_9947.png

Downloads & links

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Map

FileDownload / File: emd_9947.map.gz / Format: CCP4 / Size: 4.9 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationNone
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
1.1 Å/pix.
x 98 pix.
= 107.8 Å		1.1 Å/pix.
x 117 pix.
= 128.7 Å		1.1 Å/pix.
x 113 pix.
= 124.3 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

generated in cubic-lattice coordinate

Voxel sizeX=Y=Z: 1.1 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.0135 / Movie #1: 0.0135
Minimum - Maximum-0.007267787 - 0.023820158
Average (Standard dev.)0.0014931795 (±0.0038332199)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin394857
Dimensions11711398
Spacing11311798
CellA: 124.3 Å / B: 128.7 Å / C: 107.8 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z1.11.11.1
M x/y/z11311798
origin x/y/z0.0000.0000.000
length x/y/z124.300128.700107.800
α/β/γ90.00090.00090.000
MAP C/R/S123
start NC/NR/NS483957
NC/NR/NS11311798
D min/max/mean-0.0070.0240.001

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Supplemental data

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Additional map: cropped 12mer from filament, used for model alignment...

Fileemd_9947_additional.map
Annotationcropped 12mer from filament, used for model alignment and refinement, not for publication
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: #2

Fileemd_9947_half_map_1.map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: #1

Fileemd_9947_half_map_2.map
Projections & Slices
AxesZYX

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Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Sample components

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Entire : ASC CARD filament

EntireName: ASC CARD filament
Components
  • Complex: ASC CARD filament
    • Protein or peptide: Apoptosis-associated speck-like protein containing a CARD

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Supramolecule #1: ASC CARD filament

SupramoleculeName: ASC CARD filament / type: complex / ID: 1 / Parent: 0 / Macromolecule list: all
Source (natural)Organism: Homo sapiens (human)

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Macromolecule #1: Apoptosis-associated speck-like protein containing a CARD

MacromoleculeName: Apoptosis-associated speck-like protein containing a CARD
type: protein_or_peptide / ID: 1 / Number of copies: 12 / Enantiomer: LEVO
Source (natural)Organism: Homo sapiens (human)
Molecular weightTheoretical: 9.775143 KDa
Recombinant expressionOrganism: Escherichia coli K-12 (bacteria)
SequenceString:
LHFIDQHRAA LIARVTNVEW LLDALYGKVL TDEQYQAVRA EPTNPSKMRK LFSFTPAWNW TCKDLLLQAL RESQSYLVED LER

UniProtKB: Apoptosis-associated speck-like protein containing a CARD

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Experimental details

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Structure determination

Methodcryo EM
Processinghelical reconstruction
Aggregation statefilament

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Sample preparation

BufferpH: 8
GridModel: Quantifoil R2/2 / Material: COPPER / Mesh: 300 / Support film - Material: CARBON / Support film - topology: CONTINUOUS / Support film - Film thickness: 2 / Pretreatment - Type: GLOW DISCHARGE / Pretreatment - Time: 30 sec. / Pretreatment - Atmosphere: AIR
VitrificationCryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 277.15 K / Instrument: FEI VITROBOT MARK III

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Image recordingFilm or detector model: GATAN K2 SUMMIT (4k x 4k) / Number real images: 3183 / Average electron dose: 40.0 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsCalibrated defocus max: 2.5 µm / Calibrated defocus min: 1.5 µm / Illumination mode: FLOOD BEAM / Imaging mode: DARK FIELD / Cs: 2.7 mm / Nominal magnification: 115000
Sample stageSpecimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Cooling holder cryogen: NITROGEN
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Final reconstructionApplied symmetry - Helical parameters - Δz: 5.27 Å
Applied symmetry - Helical parameters - Δ&Phi: -100.614 °
Applied symmetry - Helical parameters - Axial symmetry: C1 (asymmetric)
Resolution.type: BY AUTHOR / Resolution: 4.1 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: RELION (ver. 2.0) / Number images used: 59566
Segment selectionNumber selected: 877408 / Software - Name: RELION (ver. 2.0)
Startup modelType of model: OTHER / Details: solid column
Final angle assignmentType: NOT APPLICABLE / Software - Name: RELION (ver. 2.0)
FSC plot (resolution estimation)

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Atomic model buiding 1

RefinementSpace: REAL / Protocol: RIGID BODY FIT
Output model

PDB-6k99:
Structure of ASC CARD filament

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