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- EMDB-28277: Cryo-EM structure of E. coli CsgA fibril (260 pixel box size) -

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Basic information

Entry
Database: EMDB / ID: EMD-28277
TitleCryo-EM structure of E. coli CsgA fibril (260 pixel box size)
Map dataE coli CsgA fibril (260 pixel box size) sharpened map
Sample
  • Complex: E. coli CsgA fibril
    • Protein or peptide: Major curlin subunit
KeywordsCurli / CsgA fibril / biofilm / PROTEIN FIBRIL
Function / homologyCurlin associated / Curlin associated repeat / regulation of amyloid fibril formation / single-species biofilm formation / pilus / amyloid fibril formation / cell adhesion / identical protein binding / Major curlin subunit
Function and homology information
Biological speciesEscherichia coli (E. coli)
Methodsingle particle reconstruction / cryo EM / Resolution: 3.8 Å
AuthorsBu F / Liu B
Funding support1 items
OrganizationGrant numberCountry
Not funded
CitationJournal: mBio / Year: 2024
Title: Structural insight into CsgA amyloid fibril assembly.
Authors: Fan Bu / Derek R Dee / Bin Liu /
Abstract: The discovery of functional amyloids in bacteria dates back several decades, and our understanding of the curli biogenesis system has gradually expanded over time. However, due to its high ...The discovery of functional amyloids in bacteria dates back several decades, and our understanding of the curli biogenesis system has gradually expanded over time. However, due to its high aggregation propensity and intrinsically disordered nature, CsgA, the main structural component of curli fibrils, has eluded comprehensive structural characterization. Recent advancements in cryo-electron microscopy (cryo-EM) offer a promising tool to achieve high-resolution structural insights into CsgA fibrils. In this study, we outline an approach to addressing the colloidal instability challenges associated with CsgA, achieved through engineering and electrostatic repulsion. Then, we present the cryo-EM structure of CsgA fibrils at 3.62 Å resolution. This structure provides new insights into the cross-β structure of CsgA. Additionally, our study identifies two distinct spatial arrangements within several CsgA fibrils, a 2-CsgA-fibril pair and a 3-CsgA-fibril bundle, shedding light on the intricate hierarchy of the biofilm extracellular matrix and laying the foundation for precise manipulation of CsgA-derived biomaterials.IMPORTANCEThe visualization of the architecture of CsgA amyloid fibril has been a longstanding research question, for which a high-resolution structure is still unavailable. CsgA serves as a major subunit of curli, the primary component of the extracellular matrix generated by bacteria. The support provided by this extracellular matrix enables bacterial biofilms to resist antibiotic treatment, significantly impacting human health. CsgA has been identified in members of , with pathogenic being the most well-known model system. Our novel insights into the structure of CsgA protofilaments form the basis for drug design targeting diseases associated with biofilms. Additionally, CsgA is widely researched in biomaterials due to its self-assembly characteristics. The resolved spatial arrangements of CsgA amyloids revealed in our study will further enhance the precision design of functional biomaterials. Therefore, our study uniquely contributes to the understanding of CsgA amyloids for both microbiology and material science.
History
DepositionSep 30, 2022-
Header (metadata) releaseOct 4, 2023-
Map releaseOct 4, 2023-
UpdateApr 24, 2024-
Current statusApr 24, 2024Processing site: RCSB / Status: Released

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Structure visualization

Supplemental images

emd_28277.png

Downloads & links

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Map

FileDownload / File: emd_28277.map.gz / Format: CCP4 / Size: 67 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationE coli CsgA fibril (260 pixel box size) sharpened map
Voxel sizeX=Y=Z: 0.88533 Å
Density
Contour LevelBy AUTHOR: 0.1
Minimum - Maximum-0.90514445 - 1.6351101
Average (Standard dev.)0.0024691555 (±0.025865013)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions260260260
Spacing260260260
CellA=B=C: 230.18658 Å
α=β=γ: 90.0 °

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Supplemental data

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Additional map: E coli CsgA fibril (260 pixel box size) unsharpened map

Fileemd_28277_additional_1.map
AnnotationE coli CsgA fibril (260 pixel box size) unsharpened map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: E coli CsgA fibril (260 pixel box size) half A map

Fileemd_28277_half_map_1.map
AnnotationE coli CsgA fibril (260 pixel box size) half_A map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: E coli CsgA fibril (260 pixel box size) half B map

Fileemd_28277_half_map_2.map
AnnotationE coli CsgA fibril (260 pixel box size) half_B map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Sample components

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Entire : E. coli CsgA fibril

EntireName: E. coli CsgA fibril
Components
  • Complex: E. coli CsgA fibril
    • Protein or peptide: Major curlin subunit

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Supramolecule #1: E. coli CsgA fibril

SupramoleculeName: E. coli CsgA fibril / type: complex / ID: 1 / Parent: 0 / Macromolecule list: all
Source (natural)Organism: Escherichia coli (E. coli)

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Macromolecule #1: Major curlin subunit

MacromoleculeName: Major curlin subunit / type: protein_or_peptide / ID: 1 / Number of copies: 8 / Enantiomer: LEVO
Source (natural)Organism: Escherichia coli (E. coli) / Strain: K12
Molecular weightTheoretical: 14.095764 KDa
Recombinant expressionOrganism: Escherichia coli (E. coli)
SequenceString:
MGVVPQYGGG GNHGGGGNNS GPNSELNIYQ YGGGNSALAL QTDCRNSDLT ITQHGGGNGA DVGQGSDDSS IDLTQRGFGN SATLDQWNG KNSEMTVKQF GGGNGAAVDQ TASNSSVNVT QCGFGNNATA HQYHHHHHH

UniProtKB: Major curlin subunit

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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: 10.4
GridModel: Quantifoil R1.2/1.3 / Material: COPPER / Mesh: 300 / Pretreatment - Type: GLOW DISCHARGE / Pretreatment - Time: 60 sec. / Pretreatment - Atmosphere: AIR
VitrificationCryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 295 K / Instrument: FEI VITROBOT MARK IV

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsC2 aperture diameter: 100.0 µm / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELDBright-field microscopy / Cs: 2.7 mm / Nominal defocus max: 2.5 µm / Nominal defocus min: 0.75 µm / Nominal magnification: 75300
Specialist opticsEnergy filter - Name: GIF Bioquantum / Energy filter - Slit width: 20 eV
Sample stageSpecimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Cooling holder cryogen: NITROGEN
Image recordingFilm or detector model: GATAN K3 BIOQUANTUM (6k x 4k) / Digitization - Dimensions - Width: 5760 pixel / Digitization - Dimensions - Height: 4092 pixel / Number grids imaged: 1 / Number real images: 7550 / Average electron dose: 50.0 e/Å2
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Startup modelType of model: NONE
Initial angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: cryoSPARC (ver. 3.3.2)
Final angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: cryoSPARC (ver. 3.3.2)
Final reconstructionApplied symmetry - Point group: C1 (asymmetric) / Algorithm: SIMULTANEOUS ITERATIVE (SIRT) / Resolution.type: BY AUTHOR / Resolution: 3.8 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: cryoSPARC (ver. 3.3.2) / Number images used: 328931
FSC plot (resolution estimation)

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Atomic model buiding 1

RefinementSpace: REAL / Protocol: OTHER
Output model

PDB-8enr:
Cryo-EM structure of E. coli CsgA fibril (260 pixel box size)

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