+Open data
-Basic information
Entry | Database: EMDB / ID: EMD-26999 | |||||||||
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Title | CryoEM structure of the T-pilus from Agrobacterium tumefaciens | |||||||||
Map data | T-pilus | |||||||||
Sample |
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Keywords | conjugation / VirB2 / type IV secretion system / pili / STRUCTURAL PROTEIN | |||||||||
Function / homology | Conjugal transfer TrbC/type IV secretion VirB2 / TrbC/VIRB2 pilin / type IV secretion system complex / protein secretion by the type IV secretion system / cell outer membrane / Protein virB2 Function and homology information | |||||||||
Biological species | Agrobacterium tumefaciens (bacteria) / Agrobacterium fabrum (strain C58 / ATCC 33970) (bacteria) | |||||||||
Method | single particle reconstruction / cryo EM / Resolution: 3.2 Å | |||||||||
Authors | Bui KH / Black CS | |||||||||
Funding support | Canada, 1 items
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Citation | Journal: Structure / Year: 2023 Title: Cryo-EM structure of the Agrobacterium tumefaciens T-pilus reveals the importance of positive charges in the lumen. Authors: Jaafar Amro / Corbin Black / Zakaria Jemouai / Nathan Rooney / Caroline Daneault / Natalie Zeytuni / Matthieu Ruiz / Khanh Huy Bui / Christian Baron / Abstract: Agrobacterium tumefaciens is a natural genetic engineer that transfers DNA into plants, which is the most applied process for generation of genetically modified plants. DNA transfer is mediated by a ...Agrobacterium tumefaciens is a natural genetic engineer that transfers DNA into plants, which is the most applied process for generation of genetically modified plants. DNA transfer is mediated by a type IV secretion system in the cell envelope and extracellular T-pili. We here report the cryo-electron microscopic structures of the T-pilus at 3.2-Å resolution and of the plasmid pKM101-determined N-pilus at 3-Å resolution. Both pili contain a main pilus protein (VirB2 in A. tumefaciens, TraM in pKM101) and phospholipids arranged in a five-start helical assembly. They contain positively charged amino acids in the lumen, and the lipids are positively charged in the T-pilus (phosphatidylcholine) conferring overall positive charge. Mutagenesis of the lumen-exposed Arg91 in VirB2 results in protein destabilization and loss of pilus formation. Our results reveal that different phospholipids can be incorporated into type IV secretion pili and that the charge of the lumen may be of functional importance. | |||||||||
History |
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-Structure visualization
Supplemental images |
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-Downloads & links
-EMDB archive
Map data | emd_26999.map.gz | 59.6 MB | EMDB map data format | |
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Header (meta data) | emd-26999-v30.xml emd-26999.xml | 15.9 KB 15.9 KB | Display Display | EMDB header |
Images | emd_26999.png | 110.3 KB | ||
Masks | emd_26999_msk_1.map | 64 MB | Mask map | |
Filedesc metadata | emd-26999.cif.gz | 5.4 KB | ||
Others | emd_26999_half_map_1.map.gz emd_26999_half_map_2.map.gz | 59.3 MB 59.3 MB | ||
Archive directory | http://ftp.pdbj.org/pub/emdb/structures/EMD-26999 ftp://ftp.pdbj.org/pub/emdb/structures/EMD-26999 | HTTPS FTP |
-Validation report
Summary document | emd_26999_validation.pdf.gz | 1001.5 KB | Display | EMDB validaton report |
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Full document | emd_26999_full_validation.pdf.gz | 1001 KB | Display | |
Data in XML | emd_26999_validation.xml.gz | 12.5 KB | Display | |
Data in CIF | emd_26999_validation.cif.gz | 14.7 KB | Display | |
Arichive directory | https://ftp.pdbj.org/pub/emdb/validation_reports/EMD-26999 ftp://ftp.pdbj.org/pub/emdb/validation_reports/EMD-26999 | HTTPS FTP |
-Related structure data
Related structure data | 8cueMC 8cw4C M: atomic model generated by this map C: citing same article (ref.) |
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Similar structure data | Similarity search - Function & homologyF&H Search |
-Links
EMDB pages | EMDB (EBI/PDBe) / EMDataResource |
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-Map
File | Download / File: emd_26999.map.gz / Format: CCP4 / Size: 64 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES) | ||||||||||||||||||||
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Annotation | T-pilus | ||||||||||||||||||||
Voxel size | X=Y=Z: 1.09 Å | ||||||||||||||||||||
Density |
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Symmetry | Space group: 1 | ||||||||||||||||||||
Details | EMDB XML:
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-Supplemental data
-Mask #1
File | emd_26999_msk_1.map | ||||||||||||
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Projections & Slices |
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Density Histograms |
-Half map: Half map 1
File | emd_26999_half_map_1.map | ||||||||||||
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Annotation | Half map 1 | ||||||||||||
Projections & Slices |
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Density Histograms |
-Half map: Half map 2
File | emd_26999_half_map_2.map | ||||||||||||
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Annotation | Half map 2 | ||||||||||||
Projections & Slices |
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Density Histograms |
-Sample components
-Entire : T-pilus of the type IV secretion system of Agrobacterium tumefaciens.
Entire | Name: T-pilus of the type IV secretion system of Agrobacterium tumefaciens. |
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Components |
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-Supramolecule #1: T-pilus of the type IV secretion system of Agrobacterium tumefaciens.
Supramolecule | Name: T-pilus of the type IV secretion system of Agrobacterium tumefaciens. type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1 |
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Source (natural) | Organism: Agrobacterium tumefaciens (bacteria) |
Molecular weight | Theoretical: 24.7 kDa/nm |
-Macromolecule #1: Protein virB2
Macromolecule | Name: Protein virB2 / type: protein_or_peptide / ID: 1 / Number of copies: 70 / Enantiomer: LEVO |
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Source (natural) | Organism: Agrobacterium fabrum (strain C58 / ATCC 33970) (bacteria) |
Molecular weight | Theoretical: 12.326439 KDa |
Sequence | String: MRCFERYRVH LNRLSLSNAV MRMVSGYAPS VVGAMGWSIF SSGPAAAQSA GGGTDPATMV NNICTFILGP FGQSLAVLGI VAIGISWMF GRASLGLVAG VVGGIVIMFG ASFLGKTLTG GG UniProtKB: Protein virB2 |
-Macromolecule #2: 1-PALMITOYL-2-LINOLEOYL-SN-GLYCERO-3-PHOSPHOCHOLINE
Macromolecule | Name: 1-PALMITOYL-2-LINOLEOYL-SN-GLYCERO-3-PHOSPHOCHOLINE / type: ligand / ID: 2 / Number of copies: 70 / Formula: CPL |
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Molecular weight | Theoretical: 758.06 Da |
Chemical component information | ChemComp-CPL: |
-Experimental details
-Structure determination
Method | cryo EM |
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Processing | single particle reconstruction |
Aggregation state | helical array |
-Sample preparation
Buffer | pH: 7.5 |
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Grid | Model: C-flat-1.2/1.3 / Material: COPPER / Mesh: 300 / Support film - Material: CARBON / Support film - topology: HOLEY / Pretreatment - Type: GLOW DISCHARGE / Pretreatment - Time: 10 sec. / Pretreatment - Atmosphere: AIR |
Vitrification | Cryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 277 K / Instrument: FEI VITROBOT MARK IV Details: 3 uL of the sample was repeatedly applied and manually blotted three times using the multiple blotting technique prior to the Vitrobot step to increase the concentration of pili.. |
-Electron microscopy
Microscope | FEI TITAN KRIOS |
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Image recording | Film or detector model: GATAN K3 BIOQUANTUM (6k x 4k) / Average electron dose: 40.0 e/Å2 |
Electron beam | Acceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN |
Electron optics | Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Nominal defocus max: 3.0 µm / Nominal defocus min: 0.5 µm / Nominal magnification: 81000 |
Experimental equipment | Model: Titan Krios / Image courtesy: FEI Company |
-Image processing
Startup model | Type of model: NONE |
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Final reconstruction | Resolution.type: BY AUTHOR / Resolution: 3.2 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: cryoSPARC (ver. 3.1) / Number images used: 615500 |
Initial angle assignment | Type: RANDOM ASSIGNMENT / Software - Name: cryoSPARC (ver. 3.1) |
Final angle assignment | Type: MAXIMUM LIKELIHOOD / Software - Name: cryoSPARC (ver. 3.1) |
-Atomic model buiding 1
Refinement | Protocol: AB INITIO MODEL |
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Output model | PDB-8cue: |