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- PDB-7wi3: Cryo-EM structure of E.Coli FtsH-HflkC AAA protease complex -

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Basic information

Entry
Database: PDB / ID: 7wi3
TitleCryo-EM structure of E.Coli FtsH-HflkC AAA protease complex
Components
  • ATP-dependent zinc metalloprotease FtsH
  • Modulator of FtsH protease HflC
  • Modulator of FtsH protease HflK
KeywordsMEMBRANE PROTEIN / complex
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 / response to heat / ATP hydrolysis activity / proteolysis / zinc ion binding ...membrane protein complex / plasma membrane protein complex / Hydrolases; Acting on peptide bonds (peptidases); Metalloendopeptidases / ATP-dependent peptidase activity / protein catabolic process / metalloendopeptidase activity / response to heat / ATP hydrolysis activity / proteolysis / zinc ion binding / ATP binding / plasma membrane / cytosol
Similarity search - Function
HflC / HflK / Menbrane protein HflK, N-terminal / Bacterial membrane protein N terminal / : / Stomatin/HflK/HflC family / Band 7 domain / SPFH domain / Band 7 family / prohibitin homologues / Peptidase M41, FtsH extracellular ...HflC / HflK / Menbrane protein HflK, N-terminal / Bacterial membrane protein N terminal / : / Stomatin/HflK/HflC family / Band 7 domain / SPFH domain / Band 7 family / prohibitin homologues / Peptidase M41, FtsH extracellular / FtsH Extracellular / Peptidase, FtsH / Peptidase M41 / Peptidase M41-like / Peptidase family M41 / Band 7/SPFH domain superfamily / 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 / Modulator of FtsH protease HflC / Modulator of FtsH protease HflK
Similarity search - Component
Biological speciesEscherichia coli K-12 (bacteria)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 4 Å
AuthorsQiao, Z. / Gao, Y.G.
Funding support Singapore, 1items
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, 2022Deposition site: PDBJ / Processing site: PDBJ
Revision 1.0Jun 1, 2022Provider: repository / Type: Initial release
Revision 1.1Jun 8, 2022Group: Database references / Category: citation / citation_author
Item: _citation.country / _citation.journal_abbrev ..._citation.country / _citation.journal_abbrev / _citation.journal_id_CSD / _citation.journal_id_ISSN / _citation.pdbx_database_id_DOI / _citation.title / _citation.year
Revision 1.2Jun 15, 2022Group: Database references / Category: citation
Item: _citation.journal_volume / _citation.page_first ..._citation.journal_volume / _citation.page_first / _citation.page_last / _citation.pdbx_database_id_PubMed / _citation.title
Revision 1.3Jun 26, 2024Group: Data collection / Category: chem_comp_atom / chem_comp_bond / em_admin / Item: _em_admin.last_update

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

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Assembly

Deposited unit
A: Modulator of FtsH protease HflC
B: Modulator of FtsH protease HflK
C: Modulator of FtsH protease HflC
D: Modulator of FtsH protease HflC
E: ATP-dependent zinc metalloprotease FtsH
F: Modulator of FtsH protease HflK
G: Modulator of FtsH protease HflK
H: ATP-dependent zinc metalloprotease FtsH
I: ATP-dependent zinc metalloprotease FtsH
J: ATP-dependent zinc metalloprotease FtsH
K: ATP-dependent zinc metalloprotease FtsH
L: ATP-dependent zinc metalloprotease FtsH
Y: Modulator of FtsH protease HflC
a: Modulator of FtsH protease HflK
c: Modulator of FtsH protease HflC
e: Modulator of FtsH protease HflC
g: ATP-dependent zinc metalloprotease FtsH
i: Modulator of FtsH protease HflK
k: Modulator of FtsH protease HflK
m: ATP-dependent zinc metalloprotease FtsH
o: ATP-dependent zinc metalloprotease FtsH
q: ATP-dependent zinc metalloprotease FtsH
s: ATP-dependent zinc metalloprotease FtsH
u: ATP-dependent zinc metalloprotease FtsH
Z: Modulator of FtsH protease HflC
b: Modulator of FtsH protease HflK
d: Modulator of FtsH protease HflC
f: Modulator of FtsH protease HflC
h: ATP-dependent zinc metalloprotease FtsH
j: Modulator of FtsH protease HflK
l: Modulator of FtsH protease HflK
n: ATP-dependent zinc metalloprotease FtsH
p: ATP-dependent zinc metalloprotease FtsH
r: ATP-dependent zinc metalloprotease FtsH
t: ATP-dependent zinc metalloprotease FtsH
v: ATP-dependent zinc metalloprotease FtsH
M: Modulator of FtsH protease HflC
N: Modulator of FtsH protease HflK
O: Modulator of FtsH protease HflC
P: Modulator of FtsH protease HflC
Q: ATP-dependent zinc metalloprotease FtsH
R: Modulator of FtsH protease HflK
S: Modulator of FtsH protease HflK
T: ATP-dependent zinc metalloprotease FtsH
U: ATP-dependent zinc metalloprotease FtsH
V: ATP-dependent zinc metalloprotease FtsH
W: ATP-dependent zinc metalloprotease FtsH
X: ATP-dependent zinc metalloprotease FtsH


Theoretical massNumber of molelcules
Total (without water)2,698,76148
Polymers2,698,76148
Non-polymers00
Water00
1


  • Idetical with deposited unit
  • defined by author
  • Evidence: microscopy
TypeNameSymmetry operationNumber
identity operation1_5551

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Components

#1: Protein
Modulator of FtsH protease HflC


Mass: 37703.887 Da / Num. of mol.: 12
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Escherichia coli K-12 (bacteria) / Strain: K12 / Gene: hflC, hflA, b4175, JW4133 / Production host: Escherichia coli (E. coli) / References: UniProt: P0ABC3
#2: Protein
Modulator of FtsH protease HflK


Mass: 45598.793 Da / Num. of mol.: 12
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Escherichia coli K-12 (bacteria) / Strain: K12 / Gene: hflK, hflA, b4174, JW4132 / Production host: Escherichia coli (E. coli) / References: UniProt: P0ABC7
#3: Protein ...
ATP-dependent zinc metalloprotease FtsH / Cell division protease FtsH


Mass: 70797.031 Da / Num. of mol.: 24
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Escherichia coli K-12 (bacteria) / Strain: K12 / Gene: ftsH, hflB, mrsC, std, tolZ, b3178, JW3145 / Production host: Escherichia coli (E. coli)
References: UniProt: P0AAI3, Hydrolases; Acting on peptide bonds (peptidases); Metalloendopeptidases

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Experimental details

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Experiment

ExperimentMethod: ELECTRON MICROSCOPY
EM experimentAggregation state: PARTICLE / 3D reconstruction method: single particle reconstruction

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Sample preparation

ComponentName: E.Coli protein complex / Type: COMPLEX / Entity ID: all / Source: RECOMBINANT
Source (natural)Organism: Escherichia coli (strain K12) (bacteria)
Source (recombinant)Organism: Escherichia coli (E. coli)
Buffer solutionpH: 7.5
SpecimenConc.: 0.6 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
VitrificationCryogen name: ETHANE

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Electron microscopy imaging

Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company
MicroscopyModel: FEI TITAN KRIOS
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Illumination mode: OTHER
Electron lensMode: BRIGHT FIELD / Nominal defocus max: 1500 nm / Nominal defocus min: 500 nm
Image recordingElectron dose: 60 e/Å2 / Film or detector model: GATAN K3 (6k x 4k)

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Processing

Software
NameVersionClassification
phenix.real_space_refine1.18.2_3874refinement
PHENIX1.18.2_3874refinement
CTF correctionType: PHASE FLIPPING ONLY
3D reconstructionResolution: 4 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 26863 / Symmetry type: POINT
RefinementCross valid method: NONE
Stereochemistry target values: GeoStd + Monomer Library + CDL v1.2
Displacement parametersBiso mean: 106.33 Å2
Refine LS restraints
Refine-IDTypeDev idealNumber
ELECTRON MICROSCOPYf_bond_d0.010470396
ELECTRON MICROSCOPYf_angle_d1.158695129
ELECTRON MICROSCOPYf_chiral_restr0.137910792
ELECTRON MICROSCOPYf_plane_restr0.006712416
ELECTRON MICROSCOPYf_dihedral_angle_d20.879826920

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