|Entry||Database: PDB / ID: 6bfc|
|Title||Cryo-EM structure of human insulin degrading enzyme in complex with insulin|
|Keywords||HYDROLASE/HORMONE / IDE / amyloid beta / HORMONE / HYDROLASE-HORMONE complex|
|Function / homology||Peptidase M16, zinc-binding site / Insulin family signature. / Peptidase M16, N-terminal / Insulin-like / Insulin / Insulin family / Insulin, conserved site / Peptidase M16, middle/third domain / Insulin-like superfamily / Insulin/IGF/Relaxin family ...Peptidase M16, zinc-binding site / Insulin family signature. / Peptidase M16, N-terminal / Insulin-like / Insulin / Insulin family / Insulin, conserved site / Peptidase M16, middle/third domain / Insulin-like superfamily / Insulin/IGF/Relaxin family / Insulinase (Peptidase family M16) / Peptidase M16 inactive domain / Middle or third domain of peptidase_M16 / Insulinase family, zinc-binding region signature. / Regulation of gene expression in beta cells / Peptidase M16, C-terminal / Insulin processing / Synthesis, secretion, and deacylation of Ghrelin / Regulation of insulin secretion / Ub-specific processing proteases / COPI-mediated anterograde transport / PI5P, PP2A and IER3 Regulate PI3K/AKT Signaling / IRS activation / Signal attenuation / Insulin receptor signalling cascade / Signaling by Insulin receptor / Insulin receptor recycling / Peroxisomal protein import / Amyloid fiber formation / Metalloenzyme, LuxS/M16 peptidase-like / 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 / negative regulation of glycogen catabolic process / alpha-beta T cell activation / negative regulation of fatty acid metabolic process / negative regulation of feeding behavior / negative regulation of NAD(P)H oxidase activity / determination of adult lifespan / regulation of cellular amino acid metabolic process / positive regulation of respiratory burst / negative regulation of respiratory burst involved in inflammatory response / regulation of protein secretion / negative regulation of protein oligomerization / peroxisomal matrix / positive regulation of peptide hormone secretion / positive regulation of dendritic spine maintenance / positive regulation of lipid biosynthetic process / negative regulation of oxidative stress-induced intrinsic apoptotic signaling pathway / negative regulation of blood vessel diameter / regulation of transmembrane transporter activity / negative regulation of gluconeogenesis / negative regulation of protein secretion / positive regulation of protein oligomerization / cognition / negative regulation of lipid catabolic process / negative regulation of reactive oxygen species biosynthetic process / positive regulation of cellular protein metabolic process / fatty acid homeostasis / positive regulation of glycolytic process / positive regulation of glycogen biosynthetic process / regulation of protein localization to plasma membrane / positive regulation of insulin receptor signaling pathway / positive regulation of nitric oxide mediated signal transduction / endoplasmic reticulum-Golgi intermediate compartment membrane / transport vesicle / regulation of synaptic plasticity / go:2001275: / positive regulation of brown fat cell differentiation / endosome lumen / amyloid-beta metabolic process / positive regulation of protein autophosphorylation / neuron projection maintenance / insulin-like growth factor receptor binding / positive regulation of cell differentiation / negative regulation of acute inflammatory response / regulation of protein localization / positive regulation of cytokine secretion / proteolysis involved in cellular protein catabolic process / positive regulation of mitotic nuclear division / protein targeting to peroxisome / positive regulation of protein catabolic process / positive regulation of glucose import / positive regulation of long-term synaptic potentiation / activation of protein kinase B activity / negative regulation of protein catabolic process / negative regulation of proteolysis / protein heterooligomerization / hormone activity / acute-phase response / peptide binding / peroxisome|
Function and homology information
|Specimen source||Homo sapiens (human)|
|Method||ELECTRON MICROSCOPY / single particle reconstruction / cryo EM / 3.7 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 26, 2017 / Release: Dec 27, 2017|
|Structure viewer||Molecule: |
Downloads & links
A: Insulin-degrading enzyme
B: Insulin-degrading enzyme
A: Insulin-degrading enzyme
B: Insulin-degrading enzyme
Mass: 111866.484 Da / Num. of mol.: 2 / Source: (gene. exp.) Homo sapiens (human) / Gene: IDE / Production host: Escherichia coli (E. coli) / Strain (production host): BL21(DE3) / References: UniProt: P14735, insulysin
Mass: 11989.862 Da / Num. of mol.: 2 / Source: (gene. exp.) Homo sapiens (human) / Gene: INS / Production host: Escherichia coli (E. coli) / Strain (production host): BL21(DE3) / References: UniProt: P01308
|Experiment||Method: ELECTRON MICROSCOPY|
|EM experiment||Aggregation state: PARTICLE / Reconstruction method: single particle reconstruction|
|Component||Name: Insulin bound insulin degrading enzyme / Type: COMPLEX / 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: BL21(DE3)|
|Buffer solution||pH: 7.8|
|Specimen||Conc.: 0.3 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES|
|Specimen support||Grid material: COPPER / Grid mesh size: 300 / Grid type: Homemade|
|Vitrification||Instrument: HOMEMADE PLUNGER / Cryogen name: ETHANE / Humidity: 85 % / Chamber temperature: 293 kelvins / Details: Grids made using Spotiton|
-Electron microscopy imaging
Model: Titan Krios / Image courtesy: FEI Company
|Microscopy||Microscope model: FEI TITAN KRIOS|
|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|
|Image recording||Average exposure time: 10 sec. / Electron dose: 71.4 e/Å2 / Detector mode: COUNTING / Film or detector model: GATAN K2 SUMMIT (4k x 4k) / Number of grids imaged: 3 / Number of real images: 3085|
|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: 3.7 Å / Resolution method: FSC 0.143 CUT-OFF / Number of particles: 148392 / Algorithm: FOURIER SPACE / Number of class averages: 2 / Symmetry type: POINT|
|Atomic model building||Overall b value: 111.9 / Ref protocol: FLEXIBLE FIT / Ref space: REAL|
|Refine LS restraints|
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