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- EMDB-21225: Cryo-EM structure of PilA-N/C from Geobacter sulfurreducens -

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Basic information

Entry
Database: EMDB / ID: EMD-21225
TitleCryo-EM structure of PilA-N/C from Geobacter sulfurreducens
Map dataCryo-EM structure of PilA-N-C from Geobacter sulfurreducens
Sample
  • Complex: Polymerized hetero-dimers of PilA-N and PilA-C
    • Protein or peptide: Geopilin domain 1 protein
    • Protein or peptide: Geopilin domain 2 protein
KeywordsProtein transport / Pili / PROTEIN FIBRIL
Function / homology
Function and homology information


pilus assembly / protein secretion by the type II secretion system / type II protein secretion system complex / plasma membrane
Similarity search - Function
Bacterial general secretion pathway protein G-type pilin / : / Prokaryotic N-terminal methylation site. / Prokaryotic N-terminal methylation motif / Prokaryotic N-terminal methylation site / Pilin-like
Similarity search - Domain/homology
Geopilin domain 2 protein / Geopilin domain 1 protein
Similarity search - Component
Biological speciesGeobacter sulfurreducens (bacteria)
Methodhelical reconstruction / cryo EM / Resolution: 3.8 Å
AuthorsGu Y / Srikanth V
Funding support United States, 4 items
OrganizationGrant numberCountry
National Science Foundation (NSF, United States)1749662 United States
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)1DP2AI138259-01 United States
Department of Defense (DOD, United States)W911NF-18-2-0100 United States
National Institutes of Health/Office of the DirectorNew Innovator Award United States
CitationJournal: Nature / Year: 2021
Title: Structure of Geobacter pili reveals secretory rather than nanowire behaviour.
Authors: Yangqi Gu / Vishok Srikanth / Aldo I Salazar-Morales / Ruchi Jain / J Patrick O'Brien / Sophia M Yi / Rajesh Kumar Soni / Fadel A Samatey / Sibel Ebru Yalcin / Nikhil S Malvankar /
Abstract: Extracellular electron transfer by Geobacter species through surface appendages known as microbial nanowires is important in a range of globally important environmental phenomena, as well as for ...Extracellular electron transfer by Geobacter species through surface appendages known as microbial nanowires is important in a range of globally important environmental phenomena, as well as for applications in bio-remediation, bioenergy, biofuels and bioelectronics. Since 2005, these nanowires have been thought to be type 4 pili composed solely of the PilA-N protein. However, previous structural analyses have demonstrated that, during extracellular electron transfer, cells do not produce pili but rather nanowires made up of the cytochromes OmcS and OmcZ. Here we show that Geobacter sulfurreducens binds PilA-N to PilA-C to assemble heterodimeric pili, which remain periplasmic under nanowire-producing conditions that require extracellular electron transfer. Cryo-electron microscopy revealed that C-terminal residues of PilA-N stabilize its copolymerization with PilA-C (to form PilA-N-C) through electrostatic and hydrophobic interactions that position PilA-C along the outer surface of the filament. PilA-N-C filaments lack π-stacking of aromatic side chains and show a conductivity that is 20,000-fold lower than that of OmcZ nanowires. In contrast with surface-displayed type 4 pili, PilA-N-C filaments show structure, function and localization akin to those of type 2 secretion pseudopili. The secretion of OmcS and OmcZ nanowires is lost when pilA-N is deleted and restored when PilA-N-C filaments are reconstituted. The substitution of pilA-N with the type 4 pili of other microorganisms also causes a loss of secretion of OmcZ nanowires. As all major phyla of prokaryotes use systems similar to type 4 pili, this nanowire translocation machinery may have a widespread effect in identifying the evolution and prevalence of diverse electron-transferring microorganisms and in determining nanowire assembly architecture for designing synthetic protein nanowires.
History
DepositionJan 19, 2020-
Header (metadata) releaseFeb 12, 2020-
Map releaseJul 7, 2021-
UpdateMay 29, 2024-
Current statusMay 29, 2024Processing site: RCSB / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 0.008
  • Imaged by UCSF Chimera
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  • Surface view colored by cylindrical radius
  • Surface level: 0.008
  • Imaged by UCSF Chimera
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  • Surface view with fitted model
  • Atomic models: PDB-6vk9
  • Surface level: 0.008
  • Imaged by UCSF Chimera
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  • Simplified surface model + fitted atomic model
  • Atomic modelsPDB-6vk9
  • Imaged by Jmol
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Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

emd_21225.png

Downloads & links

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Map

FileDownload / File: emd_21225.map.gz / Format: CCP4 / Size: 244.1 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationCryo-EM structure of PilA-N-C from Geobacter sulfurreducens
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
0.82 Å/pix.
x 400 pix.
= 328.8 Å		0.82 Å/pix.
x 400 pix.
= 328.8 Å		0.82 Å/pix.
x 400 pix.
= 328.8 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 0.822 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.008 / Movie #1: 0.008
Minimum - Maximum-0.0218699 - 0.03789511
Average (Standard dev.)0.00011130225 (±0.001122764)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions400400400
Spacing400400400
CellA=B=C: 328.80002 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z0.8220.8220.822
M x/y/z400400400
origin x/y/z0.0000.0000.000
length x/y/z328.800328.800328.800
α/β/γ90.00090.00090.000
MAP C/R/S123
start NC/NR/NS000
NC/NR/NS400400400
D min/max/mean-0.0220.0380.000

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Supplemental data

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Sample components

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Entire : Polymerized hetero-dimers of PilA-N and PilA-C

EntireName: Polymerized hetero-dimers of PilA-N and PilA-C
Components
  • Complex: Polymerized hetero-dimers of PilA-N and PilA-C
    • Protein or peptide: Geopilin domain 1 protein
    • Protein or peptide: Geopilin domain 2 protein

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Supramolecule #1: Polymerized hetero-dimers of PilA-N and PilA-C

SupramoleculeName: Polymerized hetero-dimers of PilA-N and PilA-C / type: complex / ID: 1 / Parent: 0 / Macromolecule list: all
Source (natural)Organism: Geobacter sulfurreducens (bacteria)

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Macromolecule #1: Geopilin domain 1 protein

MacromoleculeName: Geopilin domain 1 protein / type: protein_or_peptide / ID: 1 / Number of copies: 16 / Enantiomer: LEVO
Source (natural)Organism: Geobacter sulfurreducens (bacteria)
Molecular weightTheoretical: 6.576502 KDa
Recombinant expressionOrganism: Geobacter sulfurreducens (bacteria)
SequenceString:
FTLIELLIVV AIIGILAAIA IPQFSAYRVK AYNSAASSDL RNLKTALESA FADDQTYPPE S

UniProtKB: Geopilin domain 1 protein

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Macromolecule #2: Geopilin domain 2 protein

MacromoleculeName: Geopilin domain 2 protein / type: protein_or_peptide / ID: 2 / Number of copies: 16 / Enantiomer: LEVO
Source (natural)Organism: Geobacter sulfurreducens (bacteria)
Molecular weightTheoretical: 10.973763 KDa
Recombinant expressionOrganism: Geobacter sulfurreducens (bacteria)
SequenceString:
AGKIPTTTMG GKDFTFKPST NVSVSYFTTN GATSTAGTVN TDYAVNTKNS SGNRVFTSTN NTSNIWYIEN DAWKGKAVSD SDVTALGTG DVGKSDFSGT EWKSQ

UniProtKB: Geopilin domain 2 protein

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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: 7
Component:
ConcentrationFormulaName
0.0075 mMNa2HPO4Disodium phosphate
0.0025 mMNaH2PO4Monosodium phosphate
GridModel: Quantifoil R2/2 / Material: COPPER / Mesh: 400 / Support film - Material: CARBON / Support film - topology: HOLEY / Pretreatment - Type: GLOW DISCHARGE / Pretreatment - Time: 45 sec. / Pretreatment - Atmosphere: AIR / Pretreatment - Pressure: 0.032 kPa
VitrificationCryogen name: ETHANE

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Specialist opticsEnergy filter - Name: GIF Quantum LS / Energy filter - Slit width: 20 eV
Image recordingFilm or detector model: GATAN K2 SUMMIT (4k x 4k) / Detector mode: SUPER-RESOLUTION / Digitization - Frames/image: 2-10 / Number grids imaged: 1 / Number real images: 7664 / Average exposure time: 0.2 sec. / Average electron dose: 2.308 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsC2 aperture diameter: 100.0 µm / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Cs: 2.7 mm / Nominal defocus max: 2.5 µm / Nominal defocus min: 0.8 µm / Nominal magnification: 165000
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 reconstructionNumber classes used: 3
Applied symmetry - Helical parameters - Δz: 10.41 Å
Applied symmetry - Helical parameters - Δ&Phi: 89.01 °
Applied symmetry - Helical parameters - Axial symmetry: C1 (asymmetric)
Algorithm: FOURIER SPACE / Resolution.type: BY AUTHOR / Resolution: 3.8 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: RELION (ver. 3.05) / Number images used: 83000
Startup modelType of model: NONE
Final angle assignmentType: NOT APPLICABLE / Software - Name: RELION (ver. 3.05)
FSC plot (resolution estimation)

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Atomic model buiding 1

RefinementSpace: REAL / Protocol: AB INITIO MODEL / Target criteria: Correlation coefficient
Output model

PDB-6vk9:
Cryo-EM structure of PilA-N/C from Geobacter sulfurreducens

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