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- EMDB-27413: Cryo-EM structure of conjugative pili from Pyrobaculum calidifontis -

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Basic information

Entry
Database: EMDB / ID: EMD-27413
TitleCryo-EM structure of conjugative pili from Pyrobaculum calidifontis
Map dataConjugative pili from Pyrobaculum calidifontis
Sample
  • Organelle or cellular component: P. calidifontis pilin protein
    • Protein or peptide: Pilin protein
  • Ligand: [(2~{S},7~{S},11~{S},15~{S},19~{R},22~{R},26~{S},30~{R},34~{R},38~{S},43~{S},47~{S},51~{S},55~{R},58~{R},62~{S},66~{R},70~{R})-38-(hydroxymethyl)-7,11,15,19,22,26,30,34,43,47,51,55,58,62,66,70-hexadecamethyl-1,4,37,40-tetraoxacyclodoheptacont-2-yl]methanol
Function / homologymembrane / Uncharacterized protein
Function and homology information
Biological speciesPyrobaculum calidifontis (archaea)
Methodhelical reconstruction / cryo EM / negative staining / Resolution: 4.1 Å
AuthorsBeltran LC / Egelman EH
Funding support United States, 1 items
OrganizationGrant numberCountry
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)GM122510 United States
CitationJournal: Nat Commun / Year: 2023
Title: Archaeal DNA-import apparatus is homologous to bacterial conjugation machinery.
Authors: Leticia C Beltran / Virginija Cvirkaite-Krupovic / Jessalyn Miller / Fengbin Wang / Mark A B Kreutzberger / Jonasz B Patkowski / Tiago R D Costa / Stefan Schouten / Ilya Levental / Vincent P ...Authors: Leticia C Beltran / Virginija Cvirkaite-Krupovic / Jessalyn Miller / Fengbin Wang / Mark A B Kreutzberger / Jonasz B Patkowski / Tiago R D Costa / Stefan Schouten / Ilya Levental / Vincent P Conticello / Edward H Egelman / Mart Krupovic /
Abstract: Conjugation is a major mechanism of horizontal gene transfer promoting the spread of antibiotic resistance among human pathogens. It involves establishing a junction between a donor and a recipient ...Conjugation is a major mechanism of horizontal gene transfer promoting the spread of antibiotic resistance among human pathogens. It involves establishing a junction between a donor and a recipient cell via an extracellular appendage known as the mating pilus. In bacteria, the conjugation machinery is encoded by plasmids or transposons and typically mediates the transfer of cognate mobile genetic elements. Much less is known about conjugation in archaea. Here, we determine atomic structures by cryo-electron microscopy of three conjugative pili, two from hyperthermophilic archaea (Aeropyrum pernix and Pyrobaculum calidifontis) and one encoded by the Ti plasmid of the bacterium Agrobacterium tumefaciens, and show that the archaeal pili are homologous to bacterial mating pili. However, the archaeal conjugation machinery, known as Ced, has been 'domesticated', that is, the genes for the conjugation machinery are encoded on the chromosome rather than on mobile genetic elements, and mediates the transfer of cellular DNA.
History
DepositionJun 22, 2022-
Header (metadata) releaseMar 22, 2023-
Map releaseMar 22, 2023-
UpdateMar 22, 2023-
Current statusMar 22, 2023Processing site: RCSB / Status: Released

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Structure visualization

Supplemental images

emd_27413.png

Downloads & links

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Map

FileDownload / File: emd_27413.map.gz / Format: CCP4 / Size: 125 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationConjugative pili from Pyrobaculum calidifontis
Voxel sizeX=Y=Z: 1.08 Å
Density
Contour LevelBy AUTHOR: 0.202
Minimum - Maximum-0.28514937 - 0.5576675
Average (Standard dev.)0.0041363686 (±0.027186424)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin-160-160-160
Dimensions320320320
Spacing320320320
CellA=B=C: 345.6 Å
α=β=γ: 90.0 °

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Supplemental data

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Half map: Conjugative pili from Pyrobaculum calidifontis

Fileemd_27413_half_map_1.map
AnnotationConjugative pili from Pyrobaculum calidifontis
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: Conjugative pili from Pyrobaculum calidifontis

Fileemd_27413_half_map_2.map
AnnotationConjugative pili from Pyrobaculum calidifontis
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Sample components

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Entire : P. calidifontis pilin protein

EntireName: P. calidifontis pilin protein
Components
  • Organelle or cellular component: P. calidifontis pilin protein
    • Protein or peptide: Pilin protein
  • Ligand: [(2~{S},7~{S},11~{S},15~{S},19~{R},22~{R},26~{S},30~{R},34~{R},38~{S},43~{S},47~{S},51~{S},55~{R},58~{R},62~{S},66~{R},70~{R})-38-(hydroxymethyl)-7,11,15,19,22,26,30,34,43,47,51,55,58,62,66,70-hexadecamethyl-1,4,37,40-tetraoxacyclodoheptacont-2-yl]methanol

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Supramolecule #1: P. calidifontis pilin protein

SupramoleculeName: P. calidifontis pilin protein / type: organelle_or_cellular_component / ID: 1 / Parent: 0 / Macromolecule list: #1
Source (natural)Organism: Pyrobaculum calidifontis (archaea)

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Macromolecule #1: Pilin protein

MacromoleculeName: Pilin protein / type: protein_or_peptide / ID: 1 / Number of copies: 20 / Enantiomer: LEVO
Source (natural)Organism: Pyrobaculum calidifontis (archaea)
Molecular weightTheoretical: 11.943107 KDa
SequenceString:
TSVEFWQNIA SGVGKWLRAI FAIAFWSSLI LLTFYAIMTQ VAPSKVFRLG ALVDLIESVK TVLLGIFVFT ASVTGIIAGV AAIANAFGA SFAVSPIDVV NALIFQPIVD MVK

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Macromolecule #2: [(2~{S},7~{S},11~{S},15~{S},19~{R},22~{R},26~{S},30~{R},34~{R},38...

MacromoleculeName: [(2~{S},7~{S},11~{S},15~{S},19~{R},22~{R},26~{S},30~{R},34~{R},38~{S},43~{S},47~{S},51~{S},55~{R},58~{R},62~{S},66~{R},70~{R})-38-(hydroxymethyl)-7,11,15,19,22,26,30,34,43,47,51,55,58,62,66,70- ...Name: [(2~{S},7~{S},11~{S},15~{S},19~{R},22~{R},26~{S},30~{R},34~{R},38~{S},43~{S},47~{S},51~{S},55~{R},58~{R},62~{S},66~{R},70~{R})-38-(hydroxymethyl)-7,11,15,19,22,26,30,34,43,47,51,55,58,62,66,70-hexadecamethyl-1,4,37,40-tetraoxacyclodoheptacont-2-yl]methanol
type: ligand / ID: 2 / Number of copies: 20 / Formula: TT0
Molecular weightTheoretical: 1.302282 KDa
Chemical component information

ChemComp-TT0:
[(2~{S},7~{S},11~{S},15~{S},19~{R},22~{R},26~{S},30~{R},34~{R},38~{S},43~{S},47~{S},51~{S},55~{R},58~{R},62~{S},66~{R},70~{R})-38-(hydroxymethyl)-7,11,15,19,22,26,30,34,43,47,51,55,58,62,66,70-hexadecamethyl-1,4,37,40-tetraoxacyclodoheptacont-2-yl]methanol

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Experimental details

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Structure determination

Methodnegative staining, cryo EM
Processinghelical reconstruction
Aggregation statefilament

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Sample preparation

BufferpH: 7.5
StainingType: NEGATIVE / Material: uranyl acetate
GridModel: EMS Lacey Carbon / Pretreatment - Type: GLOW DISCHARGE / Pretreatment - Time: 30 sec.
VitrificationCryogen name: ETHANE / Chamber humidity: 90 % / Instrument: LEICA EM GP / Details: blot for 3 seconds before plunging.

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Electron microscopy

MicroscopeFEI TITAN
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: OTHER / Imaging mode: OTHER / Nominal defocus max: 2.2 µm / Nominal defocus min: 1.2 µm
Image recordingFilm or detector model: GATAN K3 (6k x 4k) / Average electron dose: 50.0 e/Å2

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Image processing

Final angle assignmentType: NOT APPLICABLE
Final reconstructionApplied symmetry - Helical parameters - Δz: 5.001 Å
Applied symmetry - Helical parameters - Δ&Phi: 74.206 °
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: cryoSPARC (ver. 3.2) / Number images used: 71981

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