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- EMDB-7066: Cryo-EM structure of human insulin degrading enzyme in complex wi... -

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Entry
Database: EMDB / ID: 7066
TitleCryo-EM structure of human insulin degrading enzyme in complex with FAB H11 heavy chain and FAB H11 light chain
Map dataInsulin degrading enzyme in complex with insulin
SampleInsulin degrading enzyme
Function/homologyinsulysin / beta-endorphin binding / Peptidase M16, middle/third domain / insulin catabolic process / bradykinin catabolic process / ubiquitin recycling / ubiquitin-dependent protein binding / insulin metabolic process / hormone catabolic process / Middle or third domain of peptidase_M16 ...insulysin / beta-endorphin binding / Peptidase M16, middle/third domain / insulin catabolic process / bradykinin catabolic process / ubiquitin recycling / ubiquitin-dependent protein binding / insulin metabolic process / hormone catabolic process / Middle or third domain of peptidase_M16 / Insulinase family, zinc-binding region signature. / Peptidase M16, zinc-binding site / Peptidase M16, N-terminal / Peptidase M16, C-terminal / cytosolic proteasome complex / Metalloenzyme, LuxS/M16 peptidase-like / insulin binding / peroxisomal matrix / amyloid-beta clearance / determination of adult lifespan / Peptidase M16 inactive domain / positive regulation of protein oligomerization / Insulinase (Peptidase family M16) / proteolysis involved in cellular protein catabolic process / amyloid-beta metabolic process / positive regulation of protein catabolic process / peptide binding / negative regulation of proteolysis / insulin receptor signaling pathway / protein heterooligomerization / peroxisome / antigen binding / metalloendopeptidase activity / virus receptor activity / Immunoglobulin V-set domain / amyloid-beta binding / adaptive immune response / protein homotetramerization / protein homooligomerization / ATPase activity / Immunoglobulin subtype / Immunoglobulin V-set domain / Immunoglobulin/major histocompatibility complex, conserved site / Immunoglobulins and major histocompatibility complex proteins signature. / Immunoglobulin C1-set / Ub-specific processing proteases / Ig-like domain profile. / Immunoglobulin-like domain / Immunoglobulin-like domain superfamily / proteolysis / signaling receptor binding / Immunoglobulin C1-set domain / Immunoglobulin-like fold / cell surface / mitochondrion / protein homodimerization activity / zinc ion binding / extracellular space / extracellular region / ATP binding / identical protein binding / plasma membrane / nucleus / cytosol / cytoplasm / Immunoglobulin gamma-1 heavy chain / Insulin-degrading enzyme / Uncharacterized protein
Function and homology information
SourceHomo sapiens / human /
Methodsingle particle reconstruction, at 6.5 Å resolution
AuthorsZhang Z / Liang WG / Bailey LJ / Tan YZ / Wei H / Kossiakoff AA / Carragher B / Potter SC / Tang WJ
CitationJournal: 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 2 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 2 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.
Copyright: 2018, Zhang et al.
Validation ReportPDB-ID: 6b7z

SummaryFull reportAbout validation report
DateDeposition: Oct 5, 2017 / Header (metadata) release: Jan 10, 2018 / Map release: Jan 10, 2018 / Last update: Apr 11, 2018

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 0.04
  • Imaged by UCSF CHIMERA
  • Download
  • Surface view colored by radius
  • Surface level: 0.04
  • Imaged by UCSF CHIMERA
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  • Surface view with fitted model
  • Atomic models: : PDB-6b7z
  • Surface level: 0.04
  • Imaged by UCSF CHIMERA
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3D viewer
Supplemental images

Downloads & links

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Map

Fileemd_7066.map.gz (map file in CCP4 format, 131073 KB)
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
320 pix
1.07 Å/pix.
= 343.36 Å
320 pix
1.07 Å/pix.
= 343.36 Å
320 pix
1.07 Å/pix.
= 343.36 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider package.

Voxel sizeX=Y=Z: 1.073 Å
Density
Contour Level:0.04 (by author), 0.04 (movie #1):
Minimum - Maximum-0.27929044 - 0.35023555
Average (Standard dev.)0.00038742728 (0.007850708)
Details

EMDB XML:

Space Group Number1
Map Geometry
Axis orderXYZ
Dimensions320320320
Origin000
Limit319319319
Spacing320320320
CellA=B=C: 343.36 Å
α=β=γ: 90.0 deg.

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z1.0731.0731.073
M x/y/z320320320
origin x/y/z0.0000.0000.000
length x/y/z343.360343.360343.360
α/β/γ90.00090.00090.000
start NX/NY/NZ
NX/NY/NZ
MAP C/R/S123
start NC/NR/NS000
NC/NR/NS320320320
D min/max/mean-0.2790.3500.000

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Supplemental data

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Mask #1

Fileemd_7066_msk_1.map ( map file in CCP4 format, 131073 KB )
Projections & Slices
AxesZYX
Projections
Slices (1/2)
Density Histograms
Data typeImage stored as Reals
Annotation detailsInsulin degrading enzyme in complex with insulin
Space group number1

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Mask #1~

Fileemd_7066_msk_1.map
Projections & Slices
AxesZYX
Projections
Slices (1/2)
Density Histograms

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Sample components

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Entire Insulin degrading enzyme

EntireName: Insulin degrading enzyme
Details: Cryo-EM structure of human Apo insulin degrading enzyme
Number of components: 4
MassExperimental: 100 kDa

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Component #1: protein, Insulin degrading enzyme

ProteinName: Insulin degrading enzyme
Details: Cryo-EM structure of human Apo insulin degrading enzyme
Recombinant expression: No
MassExperimental: 100 kDa
SourceSpecies: Homo sapiens / human /
Source (engineered)Expression System: Escherichia coli / bacteria / / / Strain: BL

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Component #2: protein, Insulin-degrading enzyme

ProteinName: Insulin-degrading enzyme / Recombinant expression: No
MassTheoretical: 111.866484 kDa
Source (engineered)Expression System: Homo sapiens / human /

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Component #3: protein, FAB H11 heavy chain

ProteinName: FAB H11 heavy chain / Recombinant expression: No
MassTheoretical: 23.057748 kDa
Source (engineered)Expression System: Homo sapiens / human /

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Component #4: protein, FAB H11 light chain

ProteinName: FAB H11 light chain / Recombinant expression: No
MassTheoretical: 23.087609 kDa
Source (engineered)Expression System: Homo sapiens / human /

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Experimental details

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Sample preparation

Specimen stateparticle
Sample solutionSpecimen conc.: 0.3 mg/ml / pH: 7.8
Support filmThe grids are homemade lacey gold nanowire grids
VitrificationInstrument: HOMEMADE PLUNGER / Cryogen name: ETHANE / Temperature: 298 K / Humidity: 85 % / Details: The cryo grids were made using Spotiton

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Electron microscopy imaging

Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company
ImagingMicroscope: FEI TITAN KRIOS / Details: The image was collected at 20-50 degree tilt
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Electron dose: 6.8 e/Å2 / Illumination mode: FLOOD BEAM
LensMagnification: 22500 X (nominal), 46598 X (calibrated) / Cs: 2.7 mm / Imaging mode: BRIGHT FIELD / Defocus: 940.0 - 2200.0 nm
Specimen HolderModel: FEI TITAN KRIOS AUTOGRID HOLDER / Temperature: K ( 70.0 - 70.0 K)
CameraDetector: GATAN K2 (4k x 4k)

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Image acquisition

Image acquisition #1Number of digital images: 509
Image acquisition #2Number of digital images: 620 / Sampling size: 5 microns

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Image processing

ProcessingMethod: single particle reconstruction / Number of projections: 16944
3D reconstructionAlgorithm: FOURIER SPACE / Software: RELION / Resolution: 6.5 Å / Resolution method: FSC 0.143 CUT-OFF
FSC plot (resolution assessment)

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Atomic model buiding

Output model

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