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- PDB-6pev: CryoEM Plasmodium falciparum M18 aspartyl aminopeptidase -

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Basic information

Entry
Database: PDB / ID: 6pev
TitleCryoEM Plasmodium falciparum M18 aspartyl aminopeptidase
ComponentsM18 aspartyl aminopeptidase
KeywordsMETAL BINDING PROTEIN / aspartyl aminopeptidase
Function / homology
Function and homology information


aspartyl aminopeptidase / aminopeptidase activity / metallopeptidase activity / proteolysis / zinc ion binding / cytoplasm
Similarity search - Function
Aminopeptidase i, Domain 2 / Aminopeptidase i, Domain 2 / Peptidase M18 / Peptidase M18, domain 2 / Aminopeptidase I zinc metalloprotease (M18) / Zn peptidases / Aminopeptidase / Roll / 3-Layer(aba) Sandwich / Mainly Beta / Alpha Beta
Similarity search - Domain/homology
aspartyl aminopeptidase
Similarity search - Component
Biological speciesPlasmodium falciparum (malaria parasite P. falciparum)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 3.2 Å
AuthorsHo, C. / Lai, M. / Zhou, Z.H.
Funding support United States, 10items
OrganizationGrant numberCountry
National Institutes of Health/National Center for Research Resources (NIH/NCRR)R01GM071940 United States
National Institutes of Health/National Center for Research Resources (NIH/NCRR)AI094386 United States
National Institutes of Health/National Center for Research Resources (NIH/NCRR)DE025567 United States
National Institutes of Health/National Center for Research Resources (NIH/NCRR)S10RR23057 United States
National Institutes of Health/National Center for Research Resources (NIH/NCRR)U24GM116792 United States
National Science Foundation (NSF, United States)DBI-1338135 United States
National Science Foundation (NSF, United States)DMR-1548924 United States
National Institutes of Health/National Heart, Lung, and Blood Institute (NIH/NHLBI)K99/R00 HL133453 United States
National Institutes of Health/National Institute Of Allergy and Infectious Diseases (NIH/NIAID)T32 AI007323 United States
National Institutes of Health/Office of the DirectorS10OD018111 United States
CitationJournal: Nat Methods / Year: 2020
Title: Bottom-up structural proteomics: cryoEM of protein complexes enriched from the cellular milieu.
Authors: Chi-Min Ho / Xiaorun Li / Mason Lai / Thomas C Terwilliger / Josh R Beck / James Wohlschlegel / Daniel E Goldberg / Anthony W P Fitzpatrick / Z Hong Zhou /
Abstract: X-ray crystallography often requires non-native constructs involving mutations or truncations, and is challenged by membrane proteins and large multicomponent complexes. We present here a bottom-up ...X-ray crystallography often requires non-native constructs involving mutations or truncations, and is challenged by membrane proteins and large multicomponent complexes. We present here a bottom-up endogenous structural proteomics approach whereby near-atomic-resolution cryo electron microscopy (cryoEM) maps are reconstructed ab initio from unidentified protein complexes enriched directly from the endogenous cellular milieu, followed by identification and atomic modeling of the proteins. The proteins in each complex are identified using cryoID, a program we developed to identify proteins in ab initio cryoEM maps. As a proof of principle, we applied this approach to the malaria-causing parasite Plasmodium falciparum, an organism that has resisted conventional structural-biology approaches, to obtain atomic models of multiple protein complexes implicated in intraerythrocytic survival of the parasite. Our approach is broadly applicable for determining structures of undiscovered protein complexes enriched directly from endogenous sources.
History
DepositionJun 21, 2019Deposition site: RCSB / Processing site: RCSB
Revision 1.0Dec 11, 2019Provider: repository / Type: Initial release
Revision 1.1Jan 15, 2020Group: Database references / Category: citation
Item: _citation.journal_volume / _citation.page_first ..._citation.journal_volume / _citation.page_first / _citation.page_last / _citation.year
Revision 1.2Oct 23, 2024Group: Data collection / Database references ...Data collection / Database references / Derived calculations / Structure summary
Category: chem_comp_atom / chem_comp_bond ...chem_comp_atom / chem_comp_bond / database_2 / em_admin / pdbx_entry_details / pdbx_modification_feature / struct_conn
Item: _database_2.pdbx_DOI / _database_2.pdbx_database_accession ..._database_2.pdbx_DOI / _database_2.pdbx_database_accession / _em_admin.last_update / _pdbx_entry_details.has_protein_modification / _struct_conn.pdbx_dist_value / _struct_conn.ptnr1_label_atom_id / _struct_conn.ptnr2_auth_seq_id / _struct_conn.ptnr2_label_asym_id

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Structure visualization

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  • Deposited structure unit
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Structure viewerMolecule:
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Assembly

Deposited unit
A: M18 aspartyl aminopeptidase
B: M18 aspartyl aminopeptidase
C: M18 aspartyl aminopeptidase
D: M18 aspartyl aminopeptidase
E: M18 aspartyl aminopeptidase
F: M18 aspartyl aminopeptidase
G: M18 aspartyl aminopeptidase
H: M18 aspartyl aminopeptidase
I: M18 aspartyl aminopeptidase
J: M18 aspartyl aminopeptidase
K: M18 aspartyl aminopeptidase
L: M18 aspartyl aminopeptidase
hetero molecules


Theoretical massNumber of molelcules
Total (without water)790,23336
Polymers788,66312
Non-polymers1,57024
Water00
1


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

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Components

#1: Protein
M18 aspartyl aminopeptidase


Mass: 65721.930 Da / Num. of mol.: 12 / Source method: isolated from a natural source
Source: (natural) Plasmodium falciparum (isolate NF54) (eukaryote)
Strain: isolate NF54 / References: UniProt: W7K6I8
#2: Chemical...
ChemComp-ZN / ZINC ION


Mass: 65.409 Da / Num. of mol.: 24 / Source method: obtained synthetically / Formula: Zn
Has ligand of interestN
Has protein modificationY

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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: Plasmodium falciparum M18 aspartyl aminopeptidase / Type: COMPLEX / Entity ID: #1 / Source: NATURAL
Source (natural)Organism: Plasmodium falciparum (malaria parasite P. falciparum)
Buffer solutionpH: 7.4
SpecimenEmbedding 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: FLOOD BEAM
Electron lensMode: BRIGHT FIELD / Calibrated defocus min: 1500 nm / Calibrated defocus max: 4000 nm / Cs: 2.7 mm / C2 aperture diameter: 50 µm
Image recordingElectron dose: 60 e/Å2 / Detector mode: COUNTING / Film or detector model: GATAN K2 SUMMIT (4k x 4k)

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Processing

EM software
IDNameVersionCategory
4cryoSPARC1CTF correction
10cryoSPARC1initial Euler assignment
11cryoSPARC1final Euler assignment
13cryoSPARC13D reconstruction
CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
SymmetryPoint symmetry: T (tetrahedral)
3D reconstructionResolution: 3.2 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 5860 / Num. of class averages: 1 / Symmetry type: POINT

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