|Entry||Database: EMDB / ID: 9137|
|Title||Cryo-EM structure of NLRC4-CARD filament|
|Function / homology||Green fluorescent protein / NACHT domain / The IPAF inflammasome / TP53 Regulates Transcription of Caspase Activators and Caspases / Caspase recruitment domain / Leucine-rich repeat domain superfamily / P-loop containing nucleoside triphosphate hydrolase / Green fluorescent protein-related / Death-like domain superfamily / Green fluorescent protein ...Green fluorescent protein / NACHT domain / The IPAF inflammasome / TP53 Regulates Transcription of Caspase Activators and Caspases / Caspase recruitment domain / Leucine-rich repeat domain superfamily / P-loop containing nucleoside triphosphate hydrolase / Green fluorescent protein-related / Death-like domain superfamily / Green fluorescent protein / NACHT nucleoside triphosphatase / NACHT-NTPase domain profile. / CARD caspase recruitment domain profile. / CARD domain / Green fluorescent protein, GFP / pyroptosis / interleukin-1 beta secretion / IPAF inflammasome complex / detection of bacterium / activation of innate immune response / bioluminescence / generation of precursor metabolites and energy / activation of cysteine-type endopeptidase activity involved in apoptotic process / regulation of apoptotic process / protein homooligomerization / positive regulation of NF-kappaB transcription factor activity / inflammatory response / positive regulation of apoptotic process / defense response to bacterium / apoptotic process / innate immune response / magnesium ion binding / protein homodimerization activity / ATP binding / identical protein binding / cytosol / EGFP / NLR family CARD domain-containing protein 4|
Function and homology information
|Source||Homo sapiens (human) / Vaccinia virus|
|Method||helical reconstruction / cryo EM / 3.4 Å resolution|
|Authors||Zheng W / Matyszewski M / Sohn J / Egelman EH|
|Citation||Journal: J. Biol. Chem. / Year: 2018|
Title: Cryo-EM structure of the NLRC4 filament provides insights into how symmetric and asymmetric supramolecular structures drive inflammasome assembly.
Authors: Mariusz Matyszewski / Weili Zheng / Jacob Lueck / Brendan Antiochos / Edward H Egelman / Jungsan Sohn
Abstract: Inflammasomes are supramolecular signaling platforms integral to innate immune defense against invading pathogens. The NOD-like receptor (NLR) family apoptosis inhibitory protein (NAIP)·NLR family ...Inflammasomes are supramolecular signaling platforms integral to innate immune defense against invading pathogens. The NOD-like receptor (NLR) family apoptosis inhibitory protein (NAIP)·NLR family caspase-recruiting domain (CARD) domain-containing 4 (NLRC4) inflammasome recognizes intracellular bacteria and induces the polymerization of the caspase-1 protease, which in turn executes maturation of interleukin-1β (IL-1β) and pyroptosis. Several high-resolution structures of the fully assembled NAIP·NLRC4 complex are available, but these structures do not resolve the architecture of the CARD filament in atomic detail. Here, we present the cryo-EM structure of the filament assembled by the CARD of human NLRC4 (NLRC4) at 3.4 Å resolution. The structure revealed that the helical architecture of the NLRC4 filament is essentially identical to that of the downstream filament assembled by the CARD of caspase-1 (casp1), but deviates from the split washer-like assembly of the NAIP·NLRC4 oligomer. Our results suggest that architectural complementarity is a major driver for the recognition between upstream and downstream CARD assemblies in inflammasomes. Furthermore, a Monte Carlo simulation of the NLRC4 filament assembly rationalized why an (un)decameric NLRC4 oligomer is optimal for assembling the helical base of the NLRC4 filament. Together, our results explain how symmetric and asymmetric supramolecular assemblies enable high-fidelity signaling in inflammasomes.
|Validation Report||PDB-ID: 6mks|
SummaryFull reportAbout validation report
|Date||Deposition: Sep 26, 2018 / Header (metadata) release: Oct 3, 2018 / Map release: Nov 7, 2018 / Last update: Jan 9, 2019|
|Structure viewer||EM map: |
Downloads & links
|File||emd_9137.map.gz (map file in CCP4 format, 8389 KB)|
|Projections & slices|
Images are generated by Spider.
|Voxel size||X=Y=Z: 1.32 Å|
CCP4 map header:
-Entire NLRC4-CARD filament
|Entire||Name: NLRC4-CARD filament / Number of components: 2|
-Component #1: protein, NLRC4-CARD filament
|Protein||Name: NLRC4-CARD filament / Recombinant expression: No|
|Source||Species: Homo sapiens (human)|
|Source (engineered)||Expression System: Escherichia coli BL21(DE3) (bacteria) / Vector: pET21b|
-Component #2: protein, Chimera protein of NLR family CARD domain-containing pro...
|Protein||Name: Chimera protein of NLR family CARD domain-containing protein 4 and EGFP|
Number of Copies: 31 / Recombinant expression: No
|Mass||Theoretical: 38.668805 kDa|
|Source||Species: Vaccinia virus|
|Source (engineered)||Expression System: Escherichia coli BL21(DE3) (bacteria)|
|Specimen||Specimen state: filament / Method: cryo EM|
|Helical parameters||Axial symmetry: C1 (asymmetric) / Delta z: 5 Å / Delta phi: 100.6 deg.|
|Sample solution||Buffer solution: 20mM HEPES at pH 7.4, 400mM NaCl, 10% glycerol, 1mM EDTA and 1mM DTT|
|Vitrification||Cryogen name: ETHANE / Humidity: 100 %|
-Electron microscopy imaging
|Imaging||Microscope: FEI TITAN|
|Electron gun||Electron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Electron dose: 42 e/Å2 / Illumination mode: FLOOD BEAM|
|Lens||Cs: 2.7 mm / Imaging mode: BRIGHT FIELD|
|Specimen Holder||Model: OTHER|
|Camera||Detector: GATAN K2 SUMMIT (4k x 4k)|
|Image acquisition||Number of digital images: 1690|
|Processing||Method: helical reconstruction|
|3D reconstruction||Algorithm: BACK PROJECTION / Software: SPIDER / Resolution: 3.4 Å / Resolution method: FSC 0.143 CUT-OFF|
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