+Open data
-Basic information
Entry | Database: PDB / ID: 7px8 | ||||||||||||||||||
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Title | CryoEM structure of mammalian acylaminoacyl-peptidase | ||||||||||||||||||
Components | Acylamino-acid-releasing enzyme | ||||||||||||||||||
Keywords | HYDROLASE / acylaminoacyl-peptidase / tetramer / aclypeptide-hydrolase / oxidized protein hydrolase / serine-protease | ||||||||||||||||||
Function / homology | Function and homology information acylaminoacyl-peptidase / omega peptidase activity / serine-type endopeptidase activity / proteolysis / cytoplasm Similarity search - Function | ||||||||||||||||||
Biological species | Sus scrofa domesticus (domestic pig) | ||||||||||||||||||
Method | ELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 3.27 Å | ||||||||||||||||||
Authors | Kiss-Szeman, A.J. / Harmat, V. / Menyhard, D.K. / Straner, P. / Jakli, I. / Hosogi, N. / Perczel, A. | ||||||||||||||||||
Funding support | Hungary, 5items
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Citation | Journal: Chem Sci / Year: 2022 Title: Cryo-EM structure of acylpeptide hydrolase reveals substrate selection by multimerization and a multi-state serine-protease triad. Authors: Anna J Kiss-Szemán / Pál Stráner / Imre Jákli / Naoki Hosogi / Veronika Harmat / Dóra K Menyhárd / András Perczel / Abstract: The first structure of tetrameric mammalian acylaminoacyl peptidase, an enzyme that functions as an upstream regulator of the proteasome through the removal of terminal -acetylated residues from its ...The first structure of tetrameric mammalian acylaminoacyl peptidase, an enzyme that functions as an upstream regulator of the proteasome through the removal of terminal -acetylated residues from its protein substrates, was determined by cryo-EM and further elucidated by MD simulations. Self-association results in a toroid-shaped quaternary structure, guided by an amyloidogenic β-edge and unique inserts. With a Pro introduced into its central β-sheet, sufficient conformational freedom is awarded to the segment containing the catalytic Ser587 that the serine protease catalytic triad alternates between active and latent states. Active site flexibility suggests that the dual function of catalysis and substrate selection are fulfilled by a novel mechanism: substrate entrance is regulated by flexible loops creating a double-gated channel system, while binding of the substrate to the active site is required for stabilization of the catalytic apparatus - as a second filter before hydrolysis. The structure not only underlines that within the family of S9 proteases homo-multimerization acts as a crucial tool for substrate selection, but it will also allow drug design targeting of the ubiquitin-proteasome system. | ||||||||||||||||||
History |
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-Structure visualization
Structure viewer | Molecule: MolmilJmol/JSmol |
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-Downloads & links
-Download
PDBx/mmCIF format | 7px8.cif.gz | 464.3 KB | Display | PDBx/mmCIF format |
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PDB format | pdb7px8.ent.gz | 383.1 KB | Display | PDB format |
PDBx/mmJSON format | 7px8.json.gz | Tree view | PDBx/mmJSON format | |
Others | Other downloads |
-Validation report
Arichive directory | https://data.pdbj.org/pub/pdb/validation_reports/px/7px8 ftp://data.pdbj.org/pub/pdb/validation_reports/px/7px8 | HTTPS FTP |
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-Related structure data
Related structure data | 13691MC M: map data used to model this data C: citing same article (ref.) |
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Similar structure data | Similarity search - Function & homologyF&H Search |
-Links
-Assembly
Deposited unit |
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1 |
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-Components
#1: Protein | Mass: 81324.391 Da / Num. of mol.: 4 / Source method: isolated from a natural source / Source: (natural) Sus scrofa domesticus (domestic pig) / References: UniProt: P19205 |
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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 |
-Sample preparation
Component | Name: Homotetramer of acylaminoacyl-peptidase / Type: COMPLEX / Entity ID: all / Source: NATURAL |
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Source (natural) | Organism: Sus scrofa domesticus (domestic pig) / Organ: liver |
Buffer solution | pH: 7.5 |
Buffer component | Conc.: 10 mM / Name: TRIS / Formula: C4H11NO3 |
Specimen | Conc.: 6 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES |
Specimen support | Grid material: COPPER / Grid mesh size: 200 divisions/in. / Grid type: Quantifoil R1.2/1.3 |
Vitrification | Instrument: LEICA EM GP / Cryogen name: ETHANE / Humidity: 90 % / Chamber temperature: 277 K |
-Electron microscopy imaging
Microscopy | Model: JEOL CRYO ARM 300 |
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Electron gun | Electron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Illumination mode: FLOOD BEAM |
Electron lens | Mode: BRIGHT FIELD / Nominal defocus max: 3000 nm / Nominal defocus min: 1000 nm / Cs: 2.7 mm / C2 aperture diameter: 70 µm |
Specimen holder | Cryogen: NITROGEN / Specimen holder model: JEOL |
Image recording | Average exposure time: 4 sec. / Electron dose: 40 e/Å2 / Film or detector model: GATAN K3 (6k x 4k) / Num. of grids imaged: 1 / Num. of real images: 1157 |
EM imaging optics | Energyfilter name: In-column Omega Filter / Energyfilter slit width: 20 eV |
Image scans | Sampling size: 5 µm / Width: 5760 / Height: 4092 |
-Processing
Software | Name: PHENIX / Version: 1.19.2_4158: / Classification: refinement | |||||||||
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EM software |
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CTF correction | Type: PHASE FLIPPING ONLY | |||||||||
3D reconstruction | Resolution: 3.27 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 50604 / Symmetry type: POINT |