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

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Basic information

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 insulinInsulin-degrading enzyme
SampleInsulin degrading enzymeInsulin-degrading enzyme:
Insulin-degrading enzyme / FAB H11 heavy chain / FAB H11 light chain
Function / homologyPeptidase M16, C-terminal / Immunoglobulin-like domain superfamily / Immunoglobulin C1-set / Metalloenzyme, LuxS/M16 peptidase-like / Immunoglobulin/major histocompatibility complex, conserved site / Peptidase M16, N-terminal / Immunoglobulin V-set domain / Immunoglobulin-like fold / Peptidase M16, zinc-binding site / Peptidase M16, middle/third domain ...Peptidase M16, C-terminal / Immunoglobulin-like domain superfamily / Immunoglobulin C1-set / Metalloenzyme, LuxS/M16 peptidase-like / Immunoglobulin/major histocompatibility complex, conserved site / Peptidase M16, N-terminal / Immunoglobulin V-set domain / Immunoglobulin-like fold / Peptidase M16, zinc-binding site / Peptidase M16, middle/third domain / Insulinase (Peptidase family M16) / Immunoglobulin subtype / Peptidase M16 inactive domain / Immunoglobulin C1-set domain / Immunoglobulin V-set domain / Middle or third domain of peptidase_M16 / Insulinase family, zinc-binding region signature. / Immunoglobulins and major histocompatibility complex proteins signature. / Ig-like domain profile. / Ub-specific processing proteases / Peroxisomal protein import / Immunoglobulin-like domain / beta-endorphin binding / insulysin / bradykinin catabolic process / insulin catabolic process / ubiquitin recycling / insulin metabolic process / 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 / antigen binding / metalloendopeptidase activity / insulin receptor signaling pathway / virus receptor activity / amyloid-beta binding / protein homotetramerization / adaptive immune response / protein homooligomerization / ATPase activity / proteolysis / signaling receptor binding / 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 / cryo EM / 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 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.
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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Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
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.

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 enzymeInsulin-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 (E. coli) / 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

SpecimenSpecimen state: particle / Method: cryo EM
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 estimation)

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Atomic model buiding

Output model

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