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Yorodumi- EMDB-21313: Cryo-EM structure of a substrate-engaged Bam complex with alterna... -
+Open data
-Basic information
Entry | Database: EMDB / ID: EMD-21313 | |||||||||||||||
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Title | Cryo-EM structure of a substrate-engaged Bam complex with alternative cysteine-cysteine crosslink | |||||||||||||||
Map data | ||||||||||||||||
Sample |
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Biological species | Escherichia coli K-12 (bacteria) | |||||||||||||||
Method | single particle reconstruction / cryo EM / Resolution: 6.5 Å | |||||||||||||||
Authors | Tomasek D / Rawson S / Lee J / Wzorek JS / Harrison SC / Li Z / Kahne D | |||||||||||||||
Funding support | United States, 4 items
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Citation | Journal: Nature / Year: 2020 Title: Structure of a nascent membrane protein as it folds on the BAM complex. Authors: David Tomasek / Shaun Rawson / James Lee / Joseph S Wzorek / Stephen C Harrison / Zongli Li / Daniel Kahne / Abstract: Mitochondria, chloroplasts and Gram-negative bacteria are encased in a double layer of membranes. The outer membrane contains proteins with a β-barrel structure. β-Barrels are sheets of β-strands ...Mitochondria, chloroplasts and Gram-negative bacteria are encased in a double layer of membranes. The outer membrane contains proteins with a β-barrel structure. β-Barrels are sheets of β-strands wrapped into a cylinder, in which the first strand is hydrogen-bonded to the final strand. Conserved multi-subunit molecular machines fold and insert these proteins into the outer membrane. One subunit of the machines is itself a β-barrel protein that has a central role in folding other β-barrels. In Gram-negative bacteria, the β-barrel assembly machine (BAM) consists of the β-barrel protein BamA, and four lipoproteins. To understand how the BAM complex accelerates folding without using exogenous energy (for example, ATP), we trapped folding intermediates on this machine. Here we report the structure of the BAM complex of Escherichia coli folding BamA itself. The BamA catalyst forms an asymmetric hybrid β-barrel with the BamA substrate. The N-terminal edge of the BamA catalyst has an antiparallel hydrogen-bonded interface with the C-terminal edge of the BamA substrate, consistent with previous crosslinking studies; the other edges of the BamA catalyst and substrate are close to each other, but curl inward and do not pair. Six hydrogen bonds in a membrane environment make the interface between the two proteins very stable. This stability allows folding, but creates a high kinetic barrier to substrate release after folding has finished. Features at each end of the substrate overcome this barrier and promote release by stepwise exchange of hydrogen bonds. This mechanism of substrate-assisted product release explains how the BAM complex can stably associate with the substrate during folding and then turn over rapidly when folding is complete. | |||||||||||||||
History |
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-Structure visualization
Movie |
Movie viewer |
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Structure viewer | EM map: SurfViewMolmilJmol/JSmol |
Supplemental images |
-Downloads & links
-EMDB archive
Map data | emd_21313.map.gz | 110.9 MB | EMDB map data format | |
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Header (meta data) | emd-21313-v30.xml emd-21313.xml | 11 KB 11 KB | Display Display | EMDB header |
FSC (resolution estimation) | emd_21313_fsc.xml | 13.1 KB | Display | FSC data file |
Images | emd_21313.png | 100.9 KB | ||
Archive directory | http://ftp.pdbj.org/pub/emdb/structures/EMD-21313 ftp://ftp.pdbj.org/pub/emdb/structures/EMD-21313 | HTTPS FTP |
-Validation report
Summary document | emd_21313_validation.pdf.gz | 78.8 KB | Display | EMDB validaton report |
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Full document | emd_21313_full_validation.pdf.gz | 77.9 KB | Display | |
Data in XML | emd_21313_validation.xml.gz | 492 B | Display | |
Arichive directory | https://ftp.pdbj.org/pub/emdb/validation_reports/EMD-21313 ftp://ftp.pdbj.org/pub/emdb/validation_reports/EMD-21313 | HTTPS FTP |
-Related structure data
-Links
EMDB pages | EMDB (EBI/PDBe) / EMDataResource |
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-Map
File | Download / File: emd_21313.map.gz / Format: CCP4 / Size: 184 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES) | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
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Voxel size | X=Y=Z: 0.85 Å | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Density |
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Symmetry | Space group: 1 | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Details | EMDB XML:
CCP4 map header:
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-Supplemental data
-Sample components
-Entire : BamABCDE bound to substrate BamA with loop 1 deleted
Entire | Name: BamABCDE bound to substrate BamA with loop 1 deleted |
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Components |
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-Supramolecule #1: BamABCDE bound to substrate BamA with loop 1 deleted
Supramolecule | Name: BamABCDE bound to substrate BamA with loop 1 deleted / type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1 Details: contains a cysteine crosslink between S439C in BamA and E800C in the substrate |
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Source (natural) | Organism: Escherichia coli K-12 (bacteria) |
Recombinant expression | Organism: Escherichia coli BL21(DE3) (bacteria) |
-Experimental details
-Structure determination
Method | cryo EM |
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Processing | single particle reconstruction |
Aggregation state | particle |
-Sample preparation
Concentration | 5 mg/mL | ||||||||||||
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Buffer | pH: 8 Component:
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Grid | Details: unspecified | ||||||||||||
Vitrification | Cryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 298 K / Instrument: FEI VITROBOT MARK IV | ||||||||||||
Details | contains a cysteine crosslink between S439C in BamA and E800C in the substrate |
-Electron microscopy
Microscope | FEI TITAN KRIOS |
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Specialist optics | Energy filter - Name: GIF Bioquantum / Energy filter - Slit width: 25 eV |
Image recording | Film or detector model: GATAN K3 BIOQUANTUM (6k x 4k) / Average electron dose: 55.0 e/Å2 |
Electron beam | Acceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN |
Electron optics | Calibrated defocus max: 3.2 µm / Calibrated defocus min: 1.1 µm / Calibrated magnification: 58717 / Illumination mode: OTHER / Imaging mode: OTHER |
Experimental equipment | Model: Titan Krios / Image courtesy: FEI Company |