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- PDB-5fn4: Cryo-EM structure of gamma secretase in class 2 of the apo- state... -

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Basic information

Entry
Database: PDB / ID: 5fn4
TitleCryo-EM structure of gamma secretase in class 2 of the apo- state ensemble
Components
  • Gamma-secretase subunit APH-1A
  • Gamma-secretase subunit PEN-2
  • Nicastrin
  • POLY ALA CHAIN
  • Presenilin-1
KeywordsHYDROLASE
Function / homology
Function and homology information


Cajal-Retzius cell differentiation / positive regulation of L-glutamate import across plasma membrane / amyloid precursor protein biosynthetic process / positive regulation of endopeptidase activity / gamma-secretase complex / aspartic endopeptidase activity, intramembrane cleaving / positive regulation of amyloid precursor protein biosynthetic process / smooth endoplasmic reticulum calcium ion homeostasis / Noncanonical activation of NOTCH3 / protein catabolic process at postsynapse ...Cajal-Retzius cell differentiation / positive regulation of L-glutamate import across plasma membrane / amyloid precursor protein biosynthetic process / positive regulation of endopeptidase activity / gamma-secretase complex / aspartic endopeptidase activity, intramembrane cleaving / positive regulation of amyloid precursor protein biosynthetic process / smooth endoplasmic reticulum calcium ion homeostasis / Noncanonical activation of NOTCH3 / protein catabolic process at postsynapse / TGFBR3 PTM regulation / : / : / Notch receptor processing / positive regulation of coagulation / negative regulation of axonogenesis / membrane protein intracellular domain proteolysis / skin morphogenesis / short-term synaptic potentiation / central nervous system myelination / T cell activation involved in immune response / choline transport / NOTCH4 Activation and Transmission of Signal to the Nucleus / ciliary rootlet / regulation of resting membrane potential / neural retina development / myeloid dendritic cell differentiation / Regulated proteolysis of p75NTR / metanephros development / brain morphogenesis / L-glutamate import across plasma membrane / endoplasmic reticulum calcium ion homeostasis / amyloid precursor protein metabolic process / embryonic limb morphogenesis / regulation of long-term synaptic potentiation / cell fate specification / aggresome / skeletal system morphogenesis / locomotion / astrocyte activation involved in immune response / regulation of synaptic vesicle cycle / regulation of postsynapse organization / myeloid cell homeostasis / regulation of canonical Wnt signaling pathway / T cell proliferation / G protein-coupled dopamine receptor signaling pathway / Hydrolases; Acting on peptide bonds (peptidases); Aspartic endopeptidases / dorsal/ventral neural tube patterning / growth factor receptor binding / Golgi cisterna membrane / azurophil granule membrane / positive regulation of amyloid fibril formation / glutamate receptor signaling pathway / positive regulation of dendritic spine development / amyloid-beta formation / amyloid precursor protein catabolic process / adult behavior / mitochondrial transport / blood vessel development / heart looping / regulation of neuron projection development / smooth endoplasmic reticulum / cerebral cortex cell migration / membrane protein ectodomain proteolysis / positive regulation of receptor recycling / nuclear outer membrane / negative regulation of epidermal growth factor receptor signaling pathway / negative regulation of apoptotic signaling pathway / EPH-ephrin mediated repulsion of cells / neuron development / hematopoietic progenitor cell differentiation / somitogenesis / Nuclear signaling by ERBB4 / Notch signaling pathway / endopeptidase activator activity / negative regulation of ubiquitin-dependent protein catabolic process / autophagosome assembly / calcium ion homeostasis / regulation of synaptic transmission, glutamatergic / Degradation of the extracellular matrix / epithelial cell proliferation / neuron projection maintenance / rough endoplasmic reticulum / thymus development / post-embryonic development / astrocyte activation / cellular response to calcium ion / cerebellum development / NOTCH2 Activation and Transmission of Signal to the Nucleus / positive regulation of glycolytic process / NRIF signals cell death from the nucleus / Activated NOTCH1 Transmits Signal to the Nucleus / dendritic shaft / apoptotic signaling pathway / PDZ domain binding / neuron cellular homeostasis / neuromuscular junction / NOTCH3 Activation and Transmission of Signal to the Nucleus / cell-cell adhesion / protein processing
Similarity search - Function
Peptidase A22A, presenilin 1 / Gamma-secretase subunit Aph-1 / Gamma-secretase aspartyl protease complex, presenilin enhancer-2 subunit / Aph-1 protein / Presenilin enhancer-2 subunit of gamma secretase / Peptidase A22A, presenilin / Presenilin, C-terminal / Presenilin / Nicastrin / Nicastrin, small lobe ...Peptidase A22A, presenilin 1 / Gamma-secretase subunit Aph-1 / Gamma-secretase aspartyl protease complex, presenilin enhancer-2 subunit / Aph-1 protein / Presenilin enhancer-2 subunit of gamma secretase / Peptidase A22A, presenilin / Presenilin, C-terminal / Presenilin / Nicastrin / Nicastrin, small lobe / Nicastrin large lobe / Nicastrin small lobe / Presenilin/signal peptide peptidase / Presenilin, signal peptide peptidase, family
Similarity search - Domain/homology
Presenilin-1 / Nicastrin / Gamma-secretase subunit APH-1A / Gamma-secretase subunit PEN-2
Similarity search - Component
Biological speciesHomo sapiens (human)
HOMO SAPIENS (human)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 4 Å
AuthorsBai, X.C. / Rajendra, E. / Yang, G.H. / Shi, Y.G. / Scheres, S.H.W.
CitationJournal: Elife / Year: 2015
Title: Sampling the conformational space of the catalytic subunit of human γ-secretase.
Authors: Xiao-chen Bai / Eeson Rajendra / Guanghui Yang / Yigong Shi / Sjors H W Scheres /
Abstract: Human γ-secretase is an intra-membrane protease that cleaves many different substrates. Aberrant cleavage of Notch is implicated in cancer, while abnormalities in cutting amyloid precursor protein ...Human γ-secretase is an intra-membrane protease that cleaves many different substrates. Aberrant cleavage of Notch is implicated in cancer, while abnormalities in cutting amyloid precursor protein lead to Alzheimer's disease. Our previous cryo-EM structure of γ-secretase revealed considerable disorder in its catalytic subunit presenilin. Here, we describe an image classification procedure that characterizes molecular plasticity at the secondary structure level, and apply this method to identify three distinct conformations in our previous sample. In one of these conformations, an additional transmembrane helix is visible that cannot be attributed to the known components of γ-secretase. In addition, we present a γ-secretase structure in complex with the dipeptidic inhibitor N-[N-(3,5-difluorophenacetyl)-L-alanyl]-S-phenylglycine t-butyl ester (DAPT). Our results reveal how conformational mobility in the second and sixth transmembrane helices of presenilin is greatly reduced upon binding of DAPT or the additional helix, and form the basis for a new model of how substrate enters the transmembrane domain.
History
DepositionNov 10, 2015Deposition site: PDBE / Processing site: PDBE
Revision 1.0Dec 16, 2015Provider: repository / Type: Initial release
Revision 1.1Dec 21, 2016Group: Database references
Revision 2.0Sep 11, 2019Group: Advisory / Atomic model ...Advisory / Atomic model / Data collection / Database references / Source and taxonomy / Structure summary
Category: atom_site / entity ...atom_site / entity / entity_name_com / entity_src_gen / pdbx_poly_seq_scheme / pdbx_unobs_or_zero_occ_residues / struct_ref
Item: _atom_site.label_seq_id / _entity.pdbx_description ..._atom_site.label_seq_id / _entity.pdbx_description / _entity_src_gen.gene_src_common_name / _entity_src_gen.pdbx_beg_seq_num / _entity_src_gen.pdbx_end_seq_num / _entity_src_gen.pdbx_gene_src_gene / _entity_src_gen.pdbx_gene_src_scientific_name / _entity_src_gen.pdbx_seq_type / _pdbx_poly_seq_scheme.auth_mon_id / _pdbx_poly_seq_scheme.auth_seq_num / _pdbx_poly_seq_scheme.pdb_ins_code / _pdbx_poly_seq_scheme.pdb_mon_id / _pdbx_poly_seq_scheme.pdb_seq_num / _pdbx_unobs_or_zero_occ_residues.PDB_ins_code / _pdbx_unobs_or_zero_occ_residues.auth_comp_id / _pdbx_unobs_or_zero_occ_residues.auth_seq_id / _pdbx_unobs_or_zero_occ_residues.label_comp_id / _pdbx_unobs_or_zero_occ_residues.label_seq_id / _struct_ref.pdbx_align_begin / _struct_ref.pdbx_seq_one_letter_code
Revision 2.1Oct 23, 2024Group: Data collection / Database references / Structure summary
Category: chem_comp_atom / chem_comp_bond ...chem_comp_atom / chem_comp_bond / database_2 / pdbx_entry_details / pdbx_modification_feature
Item: _database_2.pdbx_DOI / _database_2.pdbx_database_accession

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Structure visualization

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Assembly

Deposited unit
A: Nicastrin
B: Presenilin-1
C: Gamma-secretase subunit APH-1A
D: Gamma-secretase subunit PEN-2
G: POLY ALA CHAIN


Theoretical massNumber of molelcules
Total (without water)174,0485
Polymers174,0485
Non-polymers00
Water00
1


  • Idetical with deposited unit
  • defined by author&software
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1
MethodPISA

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Components

#1: Protein Nicastrin


Mass: 78483.570 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Cell line: HEK293F / Gene: NCSTN, KIAA0253, UNQ1874/PRO4317 / Plasmid: PMLINK / Production host: HOMO SAPIENS (human) / References: UniProt: Q92542
#2: Protein Presenilin-1 / PS-1 / Protein S182


Mass: 52713.535 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Cell line: HEK293F / Gene: PSEN1, AD3, PS1, PSNL1 / Plasmid: PMLINK / Production host: HOMO SAPIENS (human)
References: UniProt: P49768, Hydrolases; Acting on peptide bonds (peptidases); Aspartic endopeptidases
#3: Protein Gamma-secretase subunit APH-1A / APH-1a / Aph-1alpha / Presenilin-stabilization factor


Mass: 29017.943 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Cell line: HEK293F / Gene: APH1A, PSF, CGI-78, UNQ579/PRO1141 / Plasmid: PMLINK / Production host: HOMO SAPIENS (human) / References: UniProt: Q96BI3
#4: Protein Gamma-secretase subunit PEN-2 / Presenilin enhancer protein 2


Mass: 12038.029 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Cell line: HEK293F / Gene: PSENEN, PEN2, MDS033 / Plasmid: PMLINK / Production host: HOMO SAPIENS (human) / References: UniProt: Q9NZ42
#5: Protein/peptide POLY ALA CHAIN


Mass: 1794.960 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) HOMO SAPIENS (human) / Cell line: HEK293F / Plasmid: PMLINK / Production host: HOMO SAPIENS (human)
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: GAMMA SECRETASE / Type: COMPLEX
Buffer solutionName: 25 MM HEPES, PH 7.4, 150 MM NACL AND AMPHIPOL A8-35 / pH: 7.4
Details: 25 MM HEPES, PH 7.4, 150 MM NACL AND AMPHIPOL A8-35
SpecimenConc.: 6 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
Specimen supportDetails: HOLEY CARBON
VitrificationInstrument: FEI VITROBOT MARK IV / Cryogen name: ETHANE / Details: LIQUID ETHANE

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Electron microscopy imaging

Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company
MicroscopyModel: FEI TITAN KRIOS / Date: Oct 25, 2014
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Illumination mode: FLOOD BEAM
Electron lensMode: BRIGHT FIELD / Nominal magnification: 81000 X / Calibrated magnification: 35714 X / Nominal defocus max: 3200 nm / Nominal defocus min: 700 nm / Cs: 2.7 mm
Specimen holderTemperature: 85 K
Image recordingElectron dose: 38 e/Å2 / Film or detector model: GATAN K2 QUANTUM (4k x 4k)
Image scansNum. digital images: 2000

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Processing

SymmetryPoint symmetry: C1 (asymmetric)
3D reconstructionResolution: 4 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 79263 / Refinement type: HALF-MAPS REFINED INDEPENDENTLY / Symmetry type: POINT
RefinementHighest resolution: 4 Å
Refinement stepCycle: LAST / Highest resolution: 4 Å
ProteinNucleic acidLigandSolventTotal
Num. atoms9776 0 0 0 9776

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