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- EMDB-32521: Cryo-EM structure of E.coli membrane protein complex -

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Basic information

Entry
Database: EMDB / ID: EMD-32521
TitleCryo-EM structure of E.coli membrane protein complex
Map data
Sample
  • Complex: E.Coli protein complex
    • Protein or peptide: ATP-dependent zinc metalloprotease FtsH
  • Ligand: PHOSPHOAMINOPHOSPHONIC ACID-ADENYLATE ESTER
  • Ligand: ZINC ION
  • Ligand: MAGNESIUM ION
Keywordscomplex / MEMBRANE PROTEIN / HYDROLASE
Function / homology
Function and homology information


membrane protein complex / plasma membrane protein complex / Hydrolases; Acting on peptide bonds (peptidases); Metalloendopeptidases / ATP-dependent peptidase activity / protein catabolic process / metalloendopeptidase activity / ATP hydrolysis activity / proteolysis / zinc ion binding / ATP binding / plasma membrane
Similarity search - Function
Peptidase M41, FtsH extracellular / FtsH Extracellular / Peptidase, FtsH / Peptidase M41 / Peptidase M41-like / Peptidase family M41 / AAA ATPase, AAA+ lid domain / AAA+ lid domain / ATPase, AAA-type, conserved site / AAA-protein family signature. ...Peptidase M41, FtsH extracellular / FtsH Extracellular / Peptidase, FtsH / Peptidase M41 / Peptidase M41-like / Peptidase family M41 / AAA ATPase, AAA+ lid domain / AAA+ lid domain / ATPase, AAA-type, conserved site / AAA-protein family signature. / ATPase family associated with various cellular activities (AAA) / ATPase, AAA-type, core / ATPases associated with a variety of cellular activities / AAA+ ATPase domain / P-loop containing nucleoside triphosphate hydrolase
Similarity search - Domain/homology
ATP-dependent zinc metalloprotease FtsH
Similarity search - Component
Biological speciesEscherichia coli (strain K12) (bacteria) / Escherichia coli K-12 (bacteria)
Methodsingle particle reconstruction / cryo EM / Resolution: 3.4 Å
AuthorsQiao Z / Gao YG
Funding support Singapore, 1 items
OrganizationGrant numberCountry
Ministry of Education (MoE, Singapore) Singapore
CitationJournal: Cell Rep / Year: 2022
Title: Cryo-EM structure of the entire FtsH-HflKC AAA protease complex.
Authors: Zhu Qiao / Tatsuhiko Yokoyama / Xin-Fu Yan / Ing Tsyr Beh / Jian Shi / Sandip Basak / Yoshinori Akiyama / Yong-Gui Gao /
Abstract: The membrane-bound AAA protease FtsH is the key player controlling protein quality in bacteria. Two single-pass membrane proteins, HflK and HflC, interact with FtsH to modulate its proteolytic ...The membrane-bound AAA protease FtsH is the key player controlling protein quality in bacteria. Two single-pass membrane proteins, HflK and HflC, interact with FtsH to modulate its proteolytic activity. Here, we present structure of the entire FtsH-HflKC complex, comprising 12 copies of both HflK and HflC, all of which interact reciprocally to form a cage, as well as four FtsH hexamers with periplasmic domains and transmembrane helices enclosed inside the cage and cytoplasmic domains situated at the base of the cage. FtsH K61/D62/S63 in the β2-β3 loop in the periplasmic domain directly interact with HflK, contributing to complex formation. Pull-down and in vivo enzymatic activity assays validate the importance of the interacting interface for FtsH-HflKC complex formation. Structural comparison with the substrate-bound human m-AAA protease AFG3L2 offers implications for the HflKC cage in modulating substrate access to FtsH. Together, our findings provide a better understanding of FtsH-type AAA protease holoenzyme assembly and regulation.
History
DepositionJan 2, 2022-
Header (metadata) releaseJun 1, 2022-
Map releaseJun 1, 2022-
UpdateJun 26, 2024-
Current statusJun 26, 2024Processing site: PDBj / Status: Released

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Structure visualization

Supplemental images

emd_32521.png

Downloads & links

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Map

FileDownload / File: emd_32521.map.gz / Format: CCP4 / Size: 15.6 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
1.37 Å/pix.
x 160 pix.
= 219.648 Å		1.37 Å/pix.
x 160 pix.
= 219.648 Å		1.37 Å/pix.
x 160 pix.
= 219.648 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 1.3728 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.172
Minimum - Maximum-0.0025919026 - 1.6172231
Average (Standard dev.)0.0043730447 (±0.04632784)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions160160160
Spacing160160160
CellA=B=C: 219.648 Å
α=β=γ: 90.0 °

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Supplemental data

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Sample components

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Entire : E.Coli protein complex

EntireName: E.Coli protein complex
Components
  • Complex: E.Coli protein complex
    • Protein or peptide: ATP-dependent zinc metalloprotease FtsH
  • Ligand: PHOSPHOAMINOPHOSPHONIC ACID-ADENYLATE ESTER
  • Ligand: ZINC ION
  • Ligand: MAGNESIUM ION

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Supramolecule #1: E.Coli protein complex

SupramoleculeName: E.Coli protein complex / type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1
Source (natural)Organism: Escherichia coli (strain K12) (bacteria)

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Macromolecule #1: ATP-dependent zinc metalloprotease FtsH

MacromoleculeName: ATP-dependent zinc metalloprotease FtsH / type: protein_or_peptide / ID: 1 / Number of copies: 6 / Enantiomer: LEVO
EC number: Hydrolases; Acting on peptide bonds (peptidases); Metalloendopeptidases
Source (natural)Organism: Escherichia coli K-12 (bacteria) / Strain: K12
Molecular weightTheoretical: 70.795055 KDa
Recombinant expressionOrganism: Escherichia coli (E. coli)
SequenceString: MAKNLILWLV IAVVLMSVFQ SFGPSESNGR KVDYSTFLQE VNNDQVREAR INGREINVTK KDSNRYTTYI PVQDPKLLDN LLTKNVKVV GEPPEEPSLL ASIFISWFPM LLLIGVWIFF MRQMQGGGGK GAMSFGKSKA RMLTEDQIKT TFADVAGCDE A KEEVAELV ...String:
MAKNLILWLV IAVVLMSVFQ SFGPSESNGR KVDYSTFLQE VNNDQVREAR INGREINVTK KDSNRYTTYI PVQDPKLLDN LLTKNVKVV GEPPEEPSLL ASIFISWFPM LLLIGVWIFF MRQMQGGGGK GAMSFGKSKA RMLTEDQIKT TFADVAGCDE A KEEVAELV EYLREPSRFQ KLGGKIPKGV LMVGPPGTGK TLLAKAIAGE AKVPFFTISG SDFVEMFVGV GASRVRDMFE QA KKAAPCI IFIDQIDAVG RQRGAGLGGG HDEREQTLNQ MLVEMDGFEG NEGIIVIAAT NRPDVLDPAL LRPGRFDRQV VVG LPDVRG REQILKVHMR RVPLAPDIDA AIIARGTPGF SGADLANLVN EAALFAARGN KRVVSMVEFE KAKDKIMMGA ERRS MVMTE AQKESTAYHQ AGHAIIGRLV PEHDPVHKVT IIPRGRALGV TFFLPEGDAI SASRQKLESQ ISTLYGGRLA EEIIY GPEH VSTGASNDIK VATNLARNMV TQWGFSEKLG PLLYAEEEGE VFLGRSVAKA KHMSDETARI IDQEVKALIE RNYNRA RQL LTDNMDILHA MKDALMKYET IDAPQIDDLM ARRDVRPPAG WEEPGASNNS GDNGSPKAPR PVDEPRTPNP GNTMSEQ LG DK

UniProtKB: ATP-dependent zinc metalloprotease FtsH

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Macromolecule #2: PHOSPHOAMINOPHOSPHONIC ACID-ADENYLATE ESTER

MacromoleculeName: PHOSPHOAMINOPHOSPHONIC ACID-ADENYLATE ESTER / type: ligand / ID: 2 / Number of copies: 6 / Formula: ANP
Molecular weightTheoretical: 506.196 Da
Chemical component information

ChemComp-ANP:
PHOSPHOAMINOPHOSPHONIC ACID-ADENYLATE ESTER / AMP-PNP, energy-carrying molecule analogue*YM

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Macromolecule #3: ZINC ION

MacromoleculeName: ZINC ION / type: ligand / ID: 3 / Number of copies: 6 / Formula: ZN
Molecular weightTheoretical: 65.409 Da

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Macromolecule #4: MAGNESIUM ION

MacromoleculeName: MAGNESIUM ION / type: ligand / ID: 4 / Number of copies: 6 / Formula: MG
Molecular weightTheoretical: 24.305 Da

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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

Concentration0.6 mg/mL
BufferpH: 7.5
VitrificationCryogen name: ETHANE

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Image recordingFilm or detector model: GATAN K3 (6k x 4k) / Average electron dose: 60.0 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: OTHER / Imaging mode: BRIGHT FIELD / Nominal defocus max: 1.5 µm / Nominal defocus min: 0.5 µm
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Startup modelType of model: OTHER
Final reconstructionResolution.type: BY AUTHOR / Resolution: 3.4 Å / Resolution method: FSC 0.143 CUT-OFF / Number images used: 32359
Initial angle assignmentType: MAXIMUM LIKELIHOOD
Final angle assignmentType: MAXIMUM LIKELIHOOD

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