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- EMDB-28754: Client-bound structure of a DegP trimer within a 12mer cage -

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Basic information

Entry
Database: EMDB / ID: EMD-28754
TitleClient-bound structure of a DegP trimer within a 12mer cage
Map dataLocal refinement cryo-EM map of a DegP trimer bound to the client hTRF1. The trimer is within a 12mer cage structure.
Sample
  • Complex: Complex of a DegP trimer and the client protein hTRF1 from a 12mer cage structure
    • Protein or peptide: Periplasmic serine endoprotease DegP
    • Protein or peptide: Periplasmic serine endoprotease DegP
    • Protein or peptide: Telomeric repeat-binding factor 1
KeywordsProtease / chaperone / hydrolase / cage / complex
Function / homology
Function and homology information


positive regulation of shelterin complex assembly / negative regulation of establishment of protein localization to telomere / negative regulation of establishment of RNA localization to telomere / negative regulation of establishment of protein-containing complex localization to telomere / negative regulation of telomere maintenance via semi-conservative replication / negative regulation of exonuclease activity / negative regulation of telomeric D-loop disassembly / meiotic telomere clustering / peptidase Do / telomeric D-loop disassembly ...positive regulation of shelterin complex assembly / negative regulation of establishment of protein localization to telomere / negative regulation of establishment of RNA localization to telomere / negative regulation of establishment of protein-containing complex localization to telomere / negative regulation of telomere maintenance via semi-conservative replication / negative regulation of exonuclease activity / negative regulation of telomeric D-loop disassembly / meiotic telomere clustering / peptidase Do / telomeric D-loop disassembly / t-circle formation / shelterin complex / Telomere C-strand synthesis initiation / double-stranded telomeric DNA binding / Telomere C-strand (Lagging Strand) Synthesis / negative regulation of telomerase activity / Processive synthesis on the C-strand of the telomere / nuclear telomere cap complex / G-rich strand telomeric DNA binding / positive regulation of telomere maintenance / Polymerase switching on the C-strand of the telomere / ankyrin repeat binding / Removal of the Flap Intermediate from the C-strand / telomere capping / DNA binding, bending / negative regulation of telomere maintenance via telomere lengthening / response to temperature stimulus / protein quality control for misfolded or incompletely synthesized proteins / telomeric DNA binding / negative regulation of DNA replication / negative regulation of telomere maintenance via telomerase / Telomere Extension By Telomerase / telomere maintenance via telomerase / chaperone-mediated protein folding / Packaging Of Telomere Ends / Recognition and association of DNA glycosylase with site containing an affected purine / Cleavage of the damaged purine / Recognition and association of DNA glycosylase with site containing an affected pyrimidine / Cleavage of the damaged pyrimidine / Inhibition of DNA recombination at telomere / Meiotic synapsis / telomere maintenance / serine-type peptidase activity / DNA Damage/Telomere Stress Induced Senescence / spindle / fibrillar center / protein folding / outer membrane-bounded periplasmic space / peptidase activity / response to heat / microtubule binding / response to oxidative stress / chromosome, telomeric region / periplasmic space / molecular adaptor activity / nuclear body / cell division / serine-type endopeptidase activity / nucleolus / protein homodimerization activity / proteolysis / DNA binding / nucleoplasm / identical protein binding / nucleus / plasma membrane / cytoplasm
Similarity search - Function
Telomeric repeat-binding factor 1/2 / Telomere repeat-binding factor, dimerisation domain superfamily / Telomere repeat-binding factor, dimerisation domain / Telomere repeat binding factor (TRF) / Peptidase S1C, Do / Peptidase S1C / Trypsin-like peptidase domain / Myb-type HTH DNA-binding domain profile. / Myb domain / Myb-like DNA-binding domain ...Telomeric repeat-binding factor 1/2 / Telomere repeat-binding factor, dimerisation domain superfamily / Telomere repeat-binding factor, dimerisation domain / Telomere repeat binding factor (TRF) / Peptidase S1C, Do / Peptidase S1C / Trypsin-like peptidase domain / Myb-type HTH DNA-binding domain profile. / Myb domain / Myb-like DNA-binding domain / SANT SWI3, ADA2, N-CoR and TFIIIB'' DNA-binding domains / SANT/Myb domain / PDZ domain / PDZ domain profile. / Domain present in PSD-95, Dlg, and ZO-1/2. / PDZ domain / PDZ superfamily / Homeobox-like domain superfamily / Peptidase S1, PA clan
Similarity search - Domain/homology
Periplasmic serine endoprotease DegP / Telomeric repeat-binding factor 1
Similarity search - Component
Biological speciesEscherichia coli (strain K12) (bacteria) / Homo sapiens (human)
Methodsingle particle reconstruction / cryo EM / Resolution: 2.6 Å
AuthorsHarkness RW / Ripstein ZA / Di Trani JM / Kay LE
Funding support Canada, 1 items
OrganizationGrant numberCountry
Canadian Institutes of Health Research (CIHR)FDN-503573 Canada
CitationJournal: J Am Chem Soc / Year: 2023
Title: Flexible Client-Dependent Cages in the Assembly Landscape of the Periplasmic Protease-Chaperone DegP.
Authors: Robert W Harkness / Zev A Ripstein / Justin M Di Trani / Lewis E Kay /
Abstract: The periplasmic protein DegP, which is implicated in virulence factor transport leading to pathogenicity, is a bi-functional protease and chaperone that helps to maintain protein homeostasis in Gram- ...The periplasmic protein DegP, which is implicated in virulence factor transport leading to pathogenicity, is a bi-functional protease and chaperone that helps to maintain protein homeostasis in Gram-negative bacteria and is essential to bacterial survival under stress conditions. To perform these functions, DegP captures clients inside cage-like structures, which we have recently shown to form through the reorganization of high-order preformed apo oligomers, consisting of trimeric building blocks, that are structurally distinct from client-bound cages. Our previous studies suggested that these apo oligomers may allow DegP to encapsulate clients of various sizes under protein folding stresses by forming ensembles that can include extremely large cage particles, but how this occurs remains an open question. To explore the relation between cage and substrate sizes, we engineered a series of DegP clients of increasing hydrodynamic radii and analyzed their influence on DegP cage formation. We used dynamic light scattering and cryogenic electron microscopy to characterize the hydrodynamic properties and structures of the DegP cages that are adopted in response to each client. We present a series of density maps and structural models that include those for novel particles of approximately 30 and 60 monomers. Key interactions between DegP trimers and the bound clients that stabilize the cage assemblies and prime the clients for catalysis are revealed. We also provide evidence that DegP can form cages which approach subcellular organelles in terms of size.
History
DepositionNov 2, 2022-
Header (metadata) releaseNov 23, 2022-
Map releaseNov 23, 2022-
UpdateJul 5, 2023-
Current statusJul 5, 2023Processing site: RCSB / Status: Released

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Structure visualization

Supplemental images

emd_28754.png

Downloads & links

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Map

FileDownload / File: emd_28754.map.gz / Format: CCP4 / Size: 64 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationLocal refinement cryo-EM map of a DegP trimer bound to the client hTRF1. The trimer is within a 12mer cage structure.
Voxel sizeX=Y=Z: 1.06 Å
Density
Contour LevelBy AUTHOR: 0.22
Minimum - Maximum-0.8003041 - 1.8670492
Average (Standard dev.)0.0006470673 (±0.053136032)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions256256256
Spacing256256256
CellA=B=C: 271.36 Å
α=β=γ: 90.0 °

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Supplemental data

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Half map: #1

Fileemd_28754_half_map_1.map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: #2

Fileemd_28754_half_map_2.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 : Complex of a DegP trimer and the client protein hTRF1 from a 12me...

EntireName: Complex of a DegP trimer and the client protein hTRF1 from a 12mer cage structure
Components
  • Complex: Complex of a DegP trimer and the client protein hTRF1 from a 12mer cage structure
    • Protein or peptide: Periplasmic serine endoprotease DegP
    • Protein or peptide: Periplasmic serine endoprotease DegP
    • Protein or peptide: Telomeric repeat-binding factor 1

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Supramolecule #1: Complex of a DegP trimer and the client protein hTRF1 from a 12me...

SupramoleculeName: Complex of a DegP trimer and the client protein hTRF1 from a 12mer cage structure
type: complex / ID: 1 / Parent: 0 / Macromolecule list: all
Source (natural)Organism: Escherichia coli (strain K12) (bacteria)

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Macromolecule #1: Periplasmic serine endoprotease DegP

MacromoleculeName: Periplasmic serine endoprotease DegP / type: protein_or_peptide / ID: 1 / Number of copies: 3 / Enantiomer: LEVO / EC number: peptidase Do
Source (natural)Organism: Escherichia coli (strain K12) (bacteria) / Strain: K12
Molecular weightTheoretical: 36.376176 KDa
Recombinant expressionOrganism: Escherichia coli (E. coli)
SequenceString: MPSLAPMLEK VMPSVVSINV EGSTTVNTPR MPRNFQQFFG DDSPFCQEGS PFQSSPFCQG GQGGNGGGQQ QKFMALGSGV IIDADKGYV VTNNHVVDNA TVIKVQLSDG RKFDAKMVGK DPRSDIALIQ IQNPKNLTAI KMADSDALRV GDYTVAIGNP F GLGETVTS ...String:
MPSLAPMLEK VMPSVVSINV EGSTTVNTPR MPRNFQQFFG DDSPFCQEGS PFQSSPFCQG GQGGNGGGQQ QKFMALGSGV IIDADKGYV VTNNHVVDNA TVIKVQLSDG RKFDAKMVGK DPRSDIALIQ IQNPKNLTAI KMADSDALRV GDYTVAIGNP F GLGETVTS GIVSALGRSG LNAENYENFI QTDAAINRGN AGGALVNLNG ELIGINTAIL APDGGNIGIG FAIPSNMVKN LT SQMVEYG QVKRGELGIM GTELNSELAK AMKVDAQRGA FVSQVLPNSS AAKAGIKAGD VITSLNGKPI SSFAALRAQV GTM PVGSKL TLGLLRDGKQ VNVNLELQQS SQ

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Macromolecule #2: Periplasmic serine endoprotease DegP

MacromoleculeName: Periplasmic serine endoprotease DegP / type: protein_or_peptide / ID: 2 / Number of copies: 3 / Enantiomer: LEVO / EC number: peptidase Do
Source (natural)Organism: Escherichia coli (strain K12) (bacteria) / Strain: K12
Molecular weightTheoretical: 7.970229 KDa
Recombinant expressionOrganism: Escherichia coli (E. coli)
SequenceString:
AEMSNKGKDQ GVVVNNVKTG TPAAQIGLKK GDVIIGANQQ AVKNIAELRK VLDSKPSVLA LNIQRGDSTI YLLMQ

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Macromolecule #3: Telomeric repeat-binding factor 1

MacromoleculeName: Telomeric repeat-binding factor 1 / type: protein_or_peptide / ID: 3 / Number of copies: 3 / Enantiomer: LEVO
Source (natural)Organism: Homo sapiens (human)
Molecular weightTheoretical: 3.417168 KDa
Recombinant expressionOrganism: Escherichia coli (E. coli)
SequenceString:
SKILLHYKFN NRTSVMLKDR WRTMKKL

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Experimental details

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Structure determination

Methodcryo EM
Processingsingle particle reconstruction
Aggregation stateparticle

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Sample preparation

BufferpH: 7
VitrificationCryogen name: ETHANE-PROPANE

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELDBright-field microscopy / Nominal defocus max: 2.0 µm / Nominal defocus min: 0.9 µm
Image recordingFilm or detector model: FEI FALCON IV (4k x 4k) / Average electron dose: 45.0 e/Å2
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Startup modelType of model: INSILICO MODEL
Initial angle assignmentType: MAXIMUM LIKELIHOOD
Final angle assignmentType: MAXIMUM LIKELIHOOD
Final reconstructionResolution.type: BY AUTHOR / Resolution: 2.6 Å / Resolution method: FSC 0.143 CUT-OFF / Number images used: 483190
FSC plot (resolution estimation)

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