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- PDB-5fn5: Cryo-EM structure of gamma secretase in class 3 of the apo- state... -

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Basic information

Entry
Database: PDB / ID: 5fn5
TitleCryo-EM structure of gamma secretase in class 3 of the apo- state ensemble
Components
  • Gamma-secretase subunit APH-1A
  • Gamma-secretase subunit PEN-2
  • Nicastrin
  • 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 / negative regulation of core promoter binding / positive regulation of endopeptidase activity / gamma-secretase complex / aspartic endopeptidase activity, intramembrane cleaving / short-term synaptic potentiation / positive regulation of amyloid precursor protein biosynthetic process / smooth endoplasmic reticulum calcium ion homeostasis ...Cajal-Retzius cell differentiation / positive regulation of L-glutamate import across plasma membrane / amyloid precursor protein biosynthetic process / negative regulation of core promoter binding / positive regulation of endopeptidase activity / gamma-secretase complex / aspartic endopeptidase activity, intramembrane cleaving / short-term synaptic potentiation / 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 / sequestering of calcium ion / Notch receptor processing / synaptic vesicle targeting / negative regulation of axonogenesis / positive regulation of coagulation / central nervous system myelination / membrane protein intracellular domain proteolysis / choline transport / skin morphogenesis / T cell activation involved in immune response / NOTCH4 Activation and Transmission of Signal to the Nucleus / dorsal/ventral neural tube patterning / ciliary rootlet / neural retina development / regulation of resting membrane potential / L-glutamate import across plasma membrane / regulation of phosphorylation / Regulated proteolysis of p75NTR / myeloid dendritic cell differentiation / metanephros development / brain morphogenesis / locomotion / endoplasmic reticulum calcium ion homeostasis / amyloid precursor protein metabolic process / regulation of synaptic vesicle cycle / regulation of long-term synaptic potentiation / regulation of postsynapse organization / embryonic limb morphogenesis / cell fate specification / astrocyte activation involved in immune response / regulation of canonical Wnt signaling pathway / myeloid cell homeostasis / aggresome / skeletal system morphogenesis / azurophil granule membrane / growth factor receptor binding / Hydrolases; Acting on peptide bonds (peptidases); Aspartic endopeptidases / glutamate receptor signaling pathway / Golgi cisterna membrane / G protein-coupled dopamine receptor signaling pathway / positive regulation of amyloid fibril formation / protein glycosylation / blood vessel development / mitochondrial transport / amyloid-beta formation / heart looping / amyloid precursor protein catabolic process / positive regulation of dendritic spine development / regulation of neuron projection development / positive regulation of receptor recycling / cerebral cortex cell migration / nuclear outer membrane / adult behavior / smooth endoplasmic reticulum / membrane protein ectodomain proteolysis / negative regulation of apoptotic signaling pathway / EPH-ephrin mediated repulsion of cells / autophagosome assembly / negative regulation of ubiquitin-dependent protein catabolic process / endopeptidase activator activity / neuron development / NOTCH2 Activation and Transmission of Signal to the Nucleus / T cell proliferation / somitogenesis / hematopoietic progenitor cell differentiation / regulation of synaptic transmission, glutamatergic / Nuclear signaling by ERBB4 / calcium ion homeostasis / NRIF signals cell death from the nucleus / Activated NOTCH1 Transmits Signal to the Nucleus / rough endoplasmic reticulum / Notch signaling pathway / Degradation of the extracellular matrix / neuron projection maintenance / astrocyte activation / NOTCH3 Activation and Transmission of Signal to the Nucleus / cellular response to calcium ion / thymus development / cerebellum development / positive regulation of glycolytic process / epithelial cell proliferation / dendritic shaft / post-embryonic development / PDZ domain binding / neuromuscular junction / apoptotic signaling pathway / Constitutive Signaling by NOTCH1 PEST Domain Mutants
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)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 4.3 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 1.2Sep 11, 2019Group: Data collection / Database references ...Data collection / Database references / Source and taxonomy / Structure summary
Category: entity / entity_name_com ...entity / entity_name_com / entity_src_gen / struct_ref
Item: _entity.pdbx_description / _entity_src_gen.gene_src_common_name ..._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 / _struct_ref.pdbx_align_begin / _struct_ref.pdbx_seq_one_letter_code
Revision 1.3Oct 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


Theoretical massNumber of molelcules
Total (without water)172,2534
Polymers172,2534
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
Details: A DRUG DAPT WAS BOUND TO GAMMA SECRETASE COMPLEX / 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
Details: A DRUG DAPT WAS BOUND TO GAMMA SECRETASE COMPLEX / 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
Details: A DRUG DAPT WAS BOUND TO GAMMA SECRETASE COMPLEX / 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
Details: A DRUG DAPT WAS BOUND TO GAMMA SECRETASE COMPLEX / Source: (gene. exp.) Homo sapiens (human) / Cell line: HEK293F / Gene: PSENEN, PEN2, MDS033 / Plasmid: PMLINK / Production host: HOMO SAPIENS (human) / References: UniProt: Q9NZ42
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: 4.3 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 66720 / Refinement type: HALF-MAPS REFINED INDEPENDENTLY / Symmetry type: POINT
RefinementHighest resolution: 4.3 Å
Refinement stepCycle: LAST / Highest resolution: 4.3 Å
ProteinNucleic acidLigandSolventTotal
Num. atoms9645 0 0 0 9645

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