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- EMDB-26616: SfSTING with c-di-GMP single fiber -

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Basic information

Entry
Database: EMDB / ID: EMD-26616
TitleSfSTING with c-di-GMP single fiber
Map dataSFSTING with c-di-GMP single fiber sharpened map
Sample
  • Complex: SfSTING with c-di-GMP single fiber
    • Protein or peptide: CD-NTase-associated protein 12
  • Ligand: 9,9'-[(2R,3R,3aS,5S,7aR,9R,10R,10aS,12S,14aR)-3,5,10,12-tetrahydroxy-5,12-dioxidooctahydro-2H,7H-difuro[3,2-d:3',2'-j][1,3,7,9,2,8]tetraoxadiphosphacyclododecine-2,9-diyl]bis(2-amino-1,9-dihydro-6H-purin-6-one)
KeywordsSTING / bacterial / filament / ANTIVIRAL PROTEIN
Function / homology
Function and homology information


NAD+ glycohydrolase / NADP+ nucleosidase activity / NAD+ nucleosidase activity / defense response to virus / nucleotide binding
Similarity search - Function
CD-NTase-associated protein 12/Pycsar effector protein, TIR domain / CAP12/Pycsar effector protein, TIR domain / Prokaryotic STING domain / Prokaryotic STING domain
Similarity search - Domain/homology
CD-NTase-associated protein 12
Similarity search - Component
Biological speciesSphingobacterium faecium (bacteria)
Methodsingle particle reconstruction / cryo EM / Resolution: 3.3 Å
AuthorsMorehouse BR / Yip MCJ / Keszei AFA / McNamara-Bordewick NK / Shao S / Kranzusch PJ
Funding support United States, 11 items
OrganizationGrant numberCountry
The Pew Charitable Trusts United States
Burroughs Wellcome Fund United States
The G. Harold and Leila Y. Mathers Foundation United States
The Mark Foundation United States
Parker Institute for Cancer Immunotherapy United States
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)1DP2GM146250 United States
The Vallee Foundation Inc. United States
David and Lucile Packard Foundation United States
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)1DP2GM137415 United States
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)F32GM133063 United States
American Heart Association287375208 United States
CitationJournal: Nature / Year: 2022
Title: Cryo-EM structure of an active bacterial TIR-STING filament complex.
Authors: Benjamin R Morehouse / Matthew C J Yip / Alexander F A Keszei / Nora K McNamara-Bordewick / Sichen Shao / Philip J Kranzusch /
Abstract: Stimulator of interferon genes (STING) is an antiviral signalling protein that is broadly conserved in both innate immunity in animals and phage defence in prokaryotes. Activation of STING requires ...Stimulator of interferon genes (STING) is an antiviral signalling protein that is broadly conserved in both innate immunity in animals and phage defence in prokaryotes. Activation of STING requires its assembly into an oligomeric filament structure through binding of a cyclic dinucleotide, but the molecular basis of STING filament assembly and extension remains unknown. Here we use cryogenic electron microscopy to determine the structure of the active Toll/interleukin-1 receptor (TIR)-STING filament complex from a Sphingobacterium faecium cyclic-oligonucleotide-based antiphage signalling system (CBASS) defence operon. Bacterial TIR-STING filament formation is driven by STING interfaces that become exposed on high-affinity recognition of the cognate cyclic dinucleotide signal c-di-GMP. Repeating dimeric STING units stack laterally head-to-head through surface interfaces, which are also essential for human STING tetramer formation and downstream immune signalling in mammals. The active bacterial TIR-STING structure reveals further cross-filament contacts that brace the assembly and coordinate packing of the associated TIR NADase effector domains at the base of the filament to drive NAD hydrolysis. STING interface and cross-filament contacts are essential for cell growth arrest in vivo and reveal a stepwise mechanism of activation whereby STING filament assembly is required for subsequent effector activation. Our results define the structural basis of STING filament formation in prokaryotic antiviral signalling.
History
DepositionApr 9, 2022-
Header (metadata) releaseJul 27, 2022-
Map releaseJul 27, 2022-
UpdateFeb 14, 2024-
Current statusFeb 14, 2024Processing site: RCSB / Status: Released

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Structure visualization

Supplemental images

emd_26616.png

Downloads & links

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Map

FileDownload / File: emd_26616.map.gz / Format: CCP4 / Size: 125 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationSFSTING with c-di-GMP single fiber sharpened map
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
0.83 Å/pix.
x 320 pix.
= 264. Å		0.83 Å/pix.
x 320 pix.
= 264. Å		0.83 Å/pix.
x 320 pix.
= 264. Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 0.825 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.02
Minimum - Maximum-0.04932436 - 0.08780806
Average (Standard dev.)0.00019898533 (±0.0023077084)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions320320320
Spacing320320320
CellA=B=C: 264.0 Å
α=β=γ: 90.0 °

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Supplemental data

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Additional map: SFSTING with c-di-GMP single fiber unsharpened map

Fileemd_26616_additional_1.map
AnnotationSFSTING with c-di-GMP single fiber unsharpened map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: SfSTING with c-di-GMP single fiber half map 1

Fileemd_26616_half_map_1.map
AnnotationSfSTING with c-di-GMP single fiber half map 1
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: SfSTING with c-di-GMP single fiber half map 2

Fileemd_26616_half_map_2.map
AnnotationSfSTING with c-di-GMP single fiber half map 2
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Sample components

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Entire : SfSTING with c-di-GMP single fiber

EntireName: SfSTING with c-di-GMP single fiber
Components
  • Complex: SfSTING with c-di-GMP single fiber
    • Protein or peptide: CD-NTase-associated protein 12
  • Ligand: 9,9'-[(2R,3R,3aS,5S,7aR,9R,10R,10aS,12S,14aR)-3,5,10,12-tetrahydroxy-5,12-dioxidooctahydro-2H,7H-difuro[3,2-d:3',2'-j][1,3,7,9,2,8]tetraoxadiphosphacyclododecine-2,9-diyl]bis(2-amino-1,9-dihydro-6H-purin-6-one)

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Supramolecule #1: SfSTING with c-di-GMP single fiber

SupramoleculeName: SfSTING with c-di-GMP single fiber / type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1
Source (natural)Organism: Sphingobacterium faecium (bacteria)

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Macromolecule #1: CD-NTase-associated protein 12

MacromoleculeName: CD-NTase-associated protein 12 / type: protein_or_peptide / ID: 1 / Number of copies: 6 / Enantiomer: LEVO / EC number: NAD+ glycohydrolase
Source (natural)Organism: Sphingobacterium faecium (bacteria)
Molecular weightTheoretical: 36.956051 KDa
Recombinant expressionOrganism: Escherichia coli (E. coli)
SequenceString: MHHHHHHGSK KRIFIGSSSE QLTILNEIVD LLGDDVECIP WTDAFALNKS GLDSLIKQTR LADYSILIAT KDDLTKQRGE SLTKPRDNV VFEFGLFLGA AGPEKCYLIA EEDTDLPTDL DGITVAKFTR NSGQYNSLDK IVESIRTHLV KIAEMSQLGL L PSTALAIG ...String:
MHHHHHHGSK KRIFIGSSSE QLTILNEIVD LLGDDVECIP WTDAFALNKS GLDSLIKQTR LADYSILIAT KDDLTKQRGE SLTKPRDNV VFEFGLFLGA AGPEKCYLIA EEDTDLPTDL DGITVAKFTR NSGQYNSLDK IVESIRTHLV KIAEMSQLGL L PSTALAIG YYNSFIKRVC EEIHGSECVE LEGKKIKVKS FRVDVVIPET LDDNGVGNFT TLYNKRYGLS KATTCTNPAL LG TRGFPFH FKVDPPDANQ ESPVDIHLLD IPSTLSTIVE SLKLYLPSNQ VGQDFDMDYL EMRELENFAK VLKYLIGRNA ATK GYVNVL TNVKL

UniProtKB: CD-NTase-associated protein 12

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Macromolecule #2: 9,9'-[(2R,3R,3aS,5S,7aR,9R,10R,10aS,12S,14aR)-3,5,10,12-tetrahydr...

MacromoleculeName: 9,9'-[(2R,3R,3aS,5S,7aR,9R,10R,10aS,12S,14aR)-3,5,10,12-tetrahydroxy-5,12-dioxidooctahydro-2H,7H-difuro[3,2-d:3',2'-j][1,3,7,9,2,8]tetraoxadiphosphacyclododecine-2,9-diyl]bis(2-amino-1,9-dihydro-6H-purin-6-one)
type: ligand / ID: 2 / Number of copies: 3 / Formula: C2E
Molecular weightTheoretical: 690.411 Da
Chemical component information

ChemComp-C2E:
9,9'-[(2R,3R,3aS,5S,7aR,9R,10R,10aS,12S,14aR)-3,5,10,12-tetrahydroxy-5,12-dioxidooctahydro-2H,7H-difuro[3,2-d:3',2'-j][1,3,7,9,2,8]tetraoxadiphosphacyclododecine-2,9-diyl]bis(2-amino-1,9-dihydro-6H-purin-6-one)

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Experimental details

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Structure determination

Methodcryo EM
Processingsingle particle reconstruction
Aggregation statefilament

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Sample preparation

BufferpH: 7.5
VitrificationCryogen name: ETHANE

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Image recordingFilm or detector model: GATAN K3 (6k x 4k) / Average electron dose: 55.5 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Nominal defocus max: 2.1 µm / Nominal defocus min: 1.2 µm
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Startup modelType of model: INSILICO MODEL
Final reconstructionResolution.type: BY AUTHOR / Resolution: 3.3 Å / Resolution method: FSC 0.143 CUT-OFF / Number images used: 277287
Initial angle assignmentType: MAXIMUM LIKELIHOOD
Final angle assignmentType: MAXIMUM LIKELIHOOD
FSC plot (resolution estimation)

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