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- PDB-7ypi: Spiral hexamer of the substrate-free Lon protease with a Y224S mu... -
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Open data
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Basic information
Entry | Database: PDB / ID: 7ypi | ||||||
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Title | Spiral hexamer of the substrate-free Lon protease with a Y224S mutation | ||||||
![]() | Lon protease | ||||||
![]() | HYDROLASE / hydrolysis / AAA proteins | ||||||
Function / homology | ![]() endopeptidase La / ATP-dependent peptidase activity / protein quality control for misfolded or incompletely synthesized proteins / cellular response to heat / sequence-specific DNA binding / serine-type endopeptidase activity / ATP hydrolysis activity / ATP binding / identical protein binding / metal ion binding / cytoplasm Similarity search - Function | ||||||
Biological species | ![]() | ||||||
Method | ELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 3.8 Å | ||||||
![]() | Li, S. / Hsieh, K.Y. / Kuo, C.I. / Lee, S.H. / Ho, M.R. / Wang, C.H. / Zhang, K. / Chang, C.I. | ||||||
Funding support | ![]()
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![]() | ![]() Title: A 5+1 assemble-to-activate mechanism of the Lon proteolytic machine. Authors: Shanshan Li / Kan-Yen Hsieh / Chiao-I Kuo / Tzu-Chi Lin / Szu-Hui Lee / Yi-Ru Chen / Chun-Hsiung Wang / Meng-Ru Ho / See-Yeun Ting / Kaiming Zhang / Chung-I Chang / ![]() ![]() Abstract: Many AAA+ (ATPases associated with diverse cellular activities) proteins function as protein or DNA remodelers by threading the substrate through the central pore of their hexameric assemblies. In ...Many AAA+ (ATPases associated with diverse cellular activities) proteins function as protein or DNA remodelers by threading the substrate through the central pore of their hexameric assemblies. In this ATP-dependent translocating state, the substrate is gripped by the pore loops of the ATPase domains arranged in a universal right-handed spiral staircase organization. However, the process by which a AAA+ protein is activated to adopt this substrate-pore-loop arrangement remains unknown. We show here, using cryo-electron microscopy (cryo-EM), that the activation process of the Lon AAA+ protease may involve a pentameric assembly and a substrate-dependent incorporation of the sixth protomer to form the substrate-pore-loop contacts seen in the translocating state. Based on the structural results, we design truncated monomeric mutants that inhibit Lon activity by binding to the native pentamer and demonstrated that expressing these monomeric mutants in Escherichia coli cells containing functional Lon elicits specific phenotypes associated with lon deficiency, including the inhibition of persister cell formation. These findings uncover a substrate-dependent assembly process for the activation of a AAA+ protein and demonstrate a targeted approach to selectively inhibit its function within cells. | ||||||
History |
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Structure visualization
Structure viewer | Molecule: ![]() ![]() |
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Downloads & links
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Download
PDBx/mmCIF format | ![]() | 773.3 KB | Display | ![]() |
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PDB format | ![]() | 654.5 KB | Display | ![]() |
PDBx/mmJSON format | ![]() | Tree view | ![]() | |
Others | ![]() |
-Validation report
Summary document | ![]() | 1.7 MB | Display | ![]() |
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Full document | ![]() | 1.8 MB | Display | |
Data in XML | ![]() | 131.8 KB | Display | |
Data in CIF | ![]() | 201.2 KB | Display | |
Arichive directory | ![]() ![]() | HTTPS FTP |
-Related structure data
Related structure data | ![]() 34001MC ![]() 7yphC ![]() 7ypjC ![]() 7ypkC ![]() 7yuhC ![]() 7yumC ![]() 7yupC ![]() 7yutC ![]() 7yuuC ![]() 7yuvC ![]() 7yuwC ![]() 7yuxC ![]() 8k3yC M: map data used to model this data C: citing same article ( |
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Similar structure data | Similarity search - Function & homology ![]() |
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Links
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Assembly
Deposited unit | ![]()
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Components
#1: Protein | Mass: 88478.492 Da / Num. of mol.: 6 / Mutation: Y224S Source method: isolated from a genetically manipulated source Source: (gene. exp.) ![]() ![]() ![]() #2: Chemical | Has ligand of interest | Y | |
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-Experimental details
-Experiment
Experiment | Method: ELECTRON MICROSCOPY |
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EM experiment | Aggregation state: PARTICLE / 3D reconstruction method: single particle reconstruction |
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Sample preparation
Component | Name: Homo-Hexameric complex of Lon protease with ATPgammaS / Type: COMPLEX / Entity ID: #1 / Source: RECOMBINANT | ||||||||||||||||||||||||||||||
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Molecular weight | Value: 0.53 MDa / Experimental value: NO | ||||||||||||||||||||||||||||||
Source (natural) | Organism: ![]() | ||||||||||||||||||||||||||||||
Source (recombinant) | Organism: ![]() ![]() | ||||||||||||||||||||||||||||||
Buffer solution | pH: 8 | ||||||||||||||||||||||||||||||
Buffer component |
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Specimen | Conc.: 0.5 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES / Details: Lon protease was incubated with ATPgammaS | ||||||||||||||||||||||||||||||
Specimen support | Grid material: COPPER / Grid mesh size: 200 divisions/in. / Grid type: Quantifoil R1.2/1.3 | ||||||||||||||||||||||||||||||
Vitrification | Cryogen name: ETHANE / Humidity: 100 % / Chamber temperature: 277 K |
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Electron microscopy imaging
Experimental equipment | ![]() Model: Titan Krios / Image courtesy: FEI Company |
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Microscopy | Model: FEI TITAN KRIOS |
Electron gun | Electron source: ![]() |
Electron lens | Mode: BRIGHT FIELD / Nominal magnification: 105000 X / Nominal defocus max: 2000 nm / Nominal defocus min: 1200 nm / Cs: 2.7 mm |
Specimen holder | Cryogen: NITROGEN |
Image recording | Average exposure time: 1.34 sec. / Electron dose: 73.8 e/Å2 / Film or detector model: GATAN K3 BIOQUANTUM (6k x 4k) / Num. of grids imaged: 2 / Num. of real images: 12982 |
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Processing
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EM software |
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CTF correction | Type: PHASE FLIPPING ONLY | |||||||||||||||||||||||||||||||||||||||||||||
Particle selection | Num. of particles selected: 4721627 | |||||||||||||||||||||||||||||||||||||||||||||
Symmetry | Point symmetry: C1 (asymmetric) | |||||||||||||||||||||||||||||||||||||||||||||
3D reconstruction | Resolution: 3.8 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 612381 / Symmetry type: POINT | |||||||||||||||||||||||||||||||||||||||||||||
Refinement | Cross valid method: NONE Stereochemistry target values: GeoStd + Monomer Library + CDL v1.2 | |||||||||||||||||||||||||||||||||||||||||||||
Displacement parameters | Biso mean: 382.03 Å2 | |||||||||||||||||||||||||||||||||||||||||||||
Refine LS restraints |
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