|Entry||Database: PDB / ID: 6b7y|
|Title||Cryo-EM structure of human insulin degrading enzyme|
|Keywords||HYDROLASE / IDE / insulin degrading enzyme / amyloid beta / BIOSYNTHETIC PROTEIN|
|Function / homology||Peptidase M16, N-terminal / Peptidase M16, zinc-binding site / Peroxisomal protein import / Ub-specific processing proteases / Insulinase family, zinc-binding region signature. / Middle or third domain of peptidase_M16 / Peptidase M16 inactive domain / Insulinase (Peptidase family M16) / Peptidase M16, middle/third domain / Metalloenzyme, LuxS/M16 peptidase-like ...Peptidase M16, N-terminal / Peptidase M16, zinc-binding site / Peroxisomal protein import / Ub-specific processing proteases / Insulinase family, zinc-binding region signature. / Middle or third domain of peptidase_M16 / Peptidase M16 inactive domain / Insulinase (Peptidase family M16) / Peptidase M16, middle/third domain / Metalloenzyme, LuxS/M16 peptidase-like / Peptidase M16, C-terminal / beta-endorphin binding / insulysin / bradykinin catabolic process / insulin catabolic process / insulin metabolic process / ubiquitin recycling / ubiquitin-dependent protein binding / hormone catabolic process / insulin binding / cytosolic proteasome complex / amyloid-beta clearance / determination of adult lifespan / peroxisomal matrix / positive regulation of protein oligomerization / amyloid-beta metabolic process / proteolysis involved in cellular protein catabolic process / protein targeting to peroxisome / positive regulation of protein catabolic process / negative regulation of proteolysis / protein heterooligomerization / peptide binding / peroxisome / metalloendopeptidase activity / insulin receptor signaling pathway / virus receptor activity / amyloid-beta binding / protein homotetramerization / protein homooligomerization / ATPase activity / proteolysis / signaling receptor binding / cell surface / mitochondrion / protein homodimerization activity / zinc ion binding / extracellular space / ATP binding / identical protein binding / plasma membrane / nucleus / cytosol / cytoplasm / Insulin-degrading enzyme|
Function and homology information
|Specimen source||Homo sapiens (human)|
|Method||ELECTRON MICROSCOPY / single particle reconstruction / cryo EM / 6.5 Å resolution|
|Authors||Liang, W.G. / Zhang, Z. / Bailey, L.J. / Kossiakoff, A.A. / Tan, Y.Z. / Wei, H. / Carragher, B. / Potter, S.C. / Tang, W.J.|
|Citation||Journal: Elife / Year: 2018|
Title: Ensemble cryoEM elucidates the mechanism of insulin capture and degradation by human insulin degrading enzyme.
Authors: Zhening Zhang / Wenguang G Liang / Lucas J Bailey / Yong Zi Tan / Hui Wei / Andrew Wang / Mara Farcasanu / Virgil A Woods / Lauren A McCord / David Lee / Weifeng Shang / Rebecca Deprez-Poulain / Benoit Deprez / David R Liu / Akiko Koide / Shohei Koide / Anthony A Kossiakoff / Sheng Li / Bridget Carragher / Clinton S Potter / Wei-Jen Tang
Abstract: Insulin degrading enzyme (IDE) plays key roles in degrading peptides vital in type two diabetes, Alzheimer's, inflammation, and other human diseases. However, the process through which IDE recognizes ...Insulin degrading enzyme (IDE) plays key roles in degrading peptides vital in type two diabetes, Alzheimer's, inflammation, and other human diseases. However, the process through which IDE recognizes peptides that tend to form amyloid fibrils remained unsolved. We used cryoEM to understand both the apo- and insulin-bound dimeric IDE states, revealing that IDE displays a large opening between the homologous ~55 kDa N- and C-terminal halves to allow selective substrate capture based on size and charge complementarity. We also used cryoEM, X-ray crystallography, SAXS, and HDX-MS to elucidate the molecular basis of how amyloidogenic peptides stabilize the disordered IDE catalytic cleft, thereby inducing selective degradation by substrate-assisted catalysis. Furthermore, our insulin-bound IDE structures explain how IDE processively degrades insulin by stochastically cutting either chain without breaking disulfide bonds. Together, our studies provide a mechanism for how IDE selectively degrades amyloidogenic peptides and offers structural insights for developing IDE-based therapies.
SummaryFull reportAbout validation report
|Date||Deposition: Oct 5, 2017 / Release: Nov 8, 2017|
|Structure viewer||Molecule: |
Downloads & links
A: Insulin-degrading enzyme
B: Insulin-degrading enzyme
Mass: 111866.484 Da / Num. of mol.: 2 / Fragment: residues 46-1011 / Source: (gene. exp.) Homo sapiens (human) / Gene: IDE / Production host: Escherichia coli (E. coli) / References: UniProt: P14735, insulysin
|Experiment||Method: ELECTRON MICROSCOPY|
|EM experiment||Aggregation state: PARTICLE / Reconstruction method: single particle reconstruction|
|Component||Name: Insulin degrading enzymeInsulin-degrading enzyme / Type: COMPLEX|
Details: Cryo-EM structure of human Apo insulin degrading enzyme
Entity ID: 1 / Source: RECOMBINANT
|Molecular weight||Value: 0.1 MDa / Experimental value: YES|
|Source (natural)||Organism: Homo sapiens (human)|
|Source (recombinant)||Organism: Escherichia coli (E. coli) / Strain: BL|
|Buffer solution||pH: 7.8|
|Specimen||Conc.: 0.3 mg/ml / Details: The sample was monodisperse / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES|
|Specimen support||Details: The grids are homemade lacey gold nanowire grids / Grid material: GOLD / Grid mesh size: 300 / Grid type: Homemade|
|Vitrification||Instrument: HOMEMADE PLUNGER / Cryogen name: ETHANE / Humidity: 85 % / Chamber temperature: 298 kelvins / Details: The cryo grids were made using Spotiton|
-Electron microscopy imaging
Model: Titan Krios / Image courtesy: FEI Company
|Microscopy||Microscope model: FEI TITAN KRIOS / Details: The image was collected at 20-50 degree tilt|
|Electron gun||Electron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Illumination mode: FLOOD BEAM|
|Electron lens||Mode: BRIGHT FIELDBright-field microscopy / Nominal magnification: 22500 / Calibrated magnification: 46598 / Nominal defocus max: 2200 nm / Nominal defocus min: 940 nm / Cs: 2.7 mm / C2 aperture diameter: 70 mm / Alignment procedure: COMA FREE|
|Specimen holder||Cryogen: NITROGEN / Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Temperature (max): 70 kelvins / Temperature (min): 70 kelvins / Residual tilt: 10 mradians|
Imaging ID: 1 / Average exposure time: 10 sec. / Detector mode: COUNTING / Film or detector model: GATAN K2 SUMMIT (4k x 4k) / Number of grids imaged: 1
|Image scans||Sampling size: 5 microns / Width: 3710 / Height: 3838 / Movie frames/image: 50 / Used frames/image: 1-50|
|Software||Name: PHENIX / Version: 1.12_2829: / Classification: refinement|
|CTF correction||Type: PHASE FLIPPING AND AMPLITUDE CORRECTION|
|Particle selection||Number of particles selected: 762283|
|Symmetry||Point symmetry: C1|
|3D reconstruction||Resolution: 6.5 Å / Resolution method: FSC 0.143 CUT-OFF / Number of particles: 24425 / Algorithm: FOURIER SPACE / Number of class averages: 1 / Symmetry type: POINT|
|Atomic model building||Overall b value: 92 / Ref protocol: FLEXIBLE FIT / Ref space: REAL|
|Refine LS restraints|
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