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- EMDB-13691: CryoEM structure of mammalian acylaminoacyl-peptidase -

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Open data


ID or keywords:

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Basic information

Entry
Database: EMDB / ID: EMD-13691
TitleCryoEM structure of mammalian acylaminoacyl-peptidase
Map data
Sample
  • Complex: Homotetramer of acylaminoacyl-peptidase
    • Protein or peptide: Acylamino-acid-releasing enzyme
Keywordsacylaminoacyl-peptidase / tetramer / aclypeptide-hydrolase / oxidized protein hydrolase / serine-protease / HYDROLASE
Function / homology
Function and homology information


acylaminoacyl-peptidase / omega peptidase activity / serine-type endopeptidase activity / proteolysis / cytoplasm
Similarity search - Function
Acylamino-acid-releasing enzyme, N-terminal domain / Acylamino-acid-releasing enzyme, N-terminal domain / Prolyl endopeptidase family serine active site. / Peptidase S9, serine active site / Peptidase S9, prolyl oligopeptidase, catalytic domain / Prolyl oligopeptidase family / Six-bladed beta-propeller, TolB-like / Alpha/Beta hydrolase fold
Similarity search - Domain/homology
Acylamino-acid-releasing enzyme
Similarity search - Component
Biological speciesSus scrofa domesticus (domestic pig)
Methodsingle particle reconstruction / cryo EM / Resolution: 3.27 Å
AuthorsKiss-Szeman AJ / Harmat V
Funding support Hungary, 5 items
OrganizationGrant numberCountry
European Regional Development FundVEKOP-2.3.2-16-2017-00014 Hungary
European Regional Development FundVEKOP-2.3.3-15-2017-00018 Hungary
Ministry of Human Capacities2018-1.2.1-NKP-2018-00005 Hungary
National Research Development and Innovation Office (NKFIH)Thematic Excellence Program 2019, Synth+ Hungary
National Research Development and Innovation Office (NKFIH)K116305 Hungary
CitationJournal: 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
DepositionOct 8, 2021-
Header (metadata) releaseMay 25, 2022-
Map releaseMay 25, 2022-
UpdateJul 17, 2024-
Current statusJul 17, 2024Processing site: PDBe / Status: Released

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Structure visualization

Supplemental images

emd_13691.png

Downloads & links

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Map

FileDownload / File: emd_13691.map.gz / Format: CCP4 / Size: 125 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Voxel sizeX=Y=Z: 0.95 Å
Density
Contour LevelBy AUTHOR: 0.012
Minimum - Maximum-0.069472544 - 0.11974157
Average (Standard dev.)0.00013778942 (±0.003119792)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions320320320
Spacing320320320
CellA=B=C: 304.0 Å
α=β=γ: 90.0 °

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Supplemental data

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Sample components

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Entire : Homotetramer of acylaminoacyl-peptidase

EntireName: Homotetramer of acylaminoacyl-peptidase
Components
  • Complex: Homotetramer of acylaminoacyl-peptidase
    • Protein or peptide: Acylamino-acid-releasing enzyme

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Supramolecule #1: Homotetramer of acylaminoacyl-peptidase

SupramoleculeName: Homotetramer of acylaminoacyl-peptidase / type: complex / ID: 1 / Parent: 0 / Macromolecule list: all
Source (natural)Organism: Sus scrofa domesticus (domestic pig) / Organ: liver

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Macromolecule #1: Acylamino-acid-releasing enzyme

MacromoleculeName: Acylamino-acid-releasing enzyme / type: protein_or_peptide / ID: 1 / Number of copies: 4 / Enantiomer: LEVO
Source (natural)Organism: Sus scrofa domesticus (domestic pig)
Molecular weightTheoretical: 81.324391 KDa
SequenceString: MERQVLLSEP EEAAALYRGL SRQPALSAAC LGPEVTTQYG GRYRTVHTEW TQRDLERMEN IRFCRQYLVF HDGDSVVFAG PAGNSVETR GELLSRESPS GTMKAVLRKA GGTGTAEEKQ FLEVWEKNRK LKSFNLSALE KHGPVYEDDC FGCLSWSHSE T HLLYVADK ...String:
MERQVLLSEP EEAAALYRGL SRQPALSAAC LGPEVTTQYG GRYRTVHTEW TQRDLERMEN IRFCRQYLVF HDGDSVVFAG PAGNSVETR GELLSRESPS GTMKAVLRKA GGTGTAEEKQ FLEVWEKNRK LKSFNLSALE KHGPVYEDDC FGCLSWSHSE T HLLYVADK KRPKAESFFQ TKALDVTGSD DEMARTKKPD QAIKGDQFLF YEDWGENMVS KSTPVLCVLD IESGNISVLE GV PESVSPG QAFWAPGDTG VVFVGWWHEP FRLGIRFCTN RRSALYYVDL TGGKCELLSD ESVAVTSPRL SPDQCRIVYL RFP SLVPHQ QCGQLCLYDW YTRVTSVVVD IVPRQLGEDF SGIYCSLLPL GCWSADSQRV VFDSPQRSRQ DLFAVDTQMG SVTS LTAGG SGGSWKLLTI DRDLMVVQFS TPSVPPSLKV GFLPPAGKEQ AVSWVSLEEA EPFPDISWSI RVLQPPPQQE HVQYA GLDF EAILLQPSNS PEKTQVPMVV MPHGGPHSSF VTAWMLFPAM LCKMGFAVLL VNYRGSTGFG QDSILSLPGN VGHQDV KDV QFAVEQVLQE EHFDAGRVAL MGGSHGGFLS CHLIGQYPET YSACVVRNPV INIASMMGST DIPDWCMVEA GFSYSSD CL PDLSVWAAML DKSPIKYAPQ VKTPLLLMLG QEDRRVPFKQ GMEYYRVLKA RNVPVRLLLY PKSTHALSEV EVESDSFM N AVLWLCTHLG S

UniProtKB: Acylamino-acid-releasing enzyme

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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

Concentration6 mg/mL
BufferpH: 7.5 / Component - Concentration: 10.0 mM / Component - Formula: C4H11NO3 / Component - Name: TRIS
GridModel: Quantifoil R1.2/1.3 / Material: COPPER / Mesh: 200 / Pretreatment - Type: GLOW DISCHARGE / Pretreatment - Time: 30 sec. / Pretreatment - Atmosphere: AIR / Pretreatment - Pressure: 0.015 kPa
VitrificationCryogen name: ETHANE / Chamber humidity: 90 % / Chamber temperature: 277 K / Instrument: LEICA EM GP

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Electron microscopy

MicroscopeJEOL CRYO ARM 300
Specialist opticsEnergy filter - Name: In-column Omega Filter / Energy filter - Slit width: 20 eV
Image recordingFilm or detector model: GATAN K3 (6k x 4k) / Digitization - Dimensions - Width: 5760 pixel / Digitization - Dimensions - Height: 4092 pixel / Number grids imaged: 1 / Number real images: 1157 / Average exposure time: 4.0 sec. / Average electron dose: 40.0 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsC2 aperture diameter: 70.0 µm / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Cs: 2.7 mm / Nominal defocus max: 3.0 µm / Nominal defocus min: 1.0 µm
Sample stageSpecimen holder model: JEOL / Cooling holder cryogen: NITROGEN

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Image processing

Startup modelType of model: NONE
Final reconstructionResolution.type: BY AUTHOR / Resolution: 3.27 Å / Resolution method: FSC 0.143 CUT-OFF / Number images used: 50604
Initial angle assignmentType: OTHER
Final angle assignmentType: OTHER

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