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- EMDB-7041: Cryo-EM structure of human insulin degrading enzyme in complex wi... -

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Entry
Database: EMDB / ID: EMD-7041
TitleCryo-EM structure of human insulin degrading enzyme in complex with insulin
Map dataThis is the sharpened map from the final, Insulin degrading enzyme in complex with insulin
Sample
  • Complex: Insulin degrading enzyme/Insulin
    • Protein or peptide: Insulin-degrading enzyme
    • Protein or peptide: Insulin
KeywordsIDE / insulin degrading enzyme / amyloid beta / HYDROLASE-HORMONE complex
Function / homology
Function and homology information


insulysin / beta-endorphin binding / ubiquitin recycling / insulin catabolic process / insulin metabolic process / amyloid-beta clearance by cellular catabolic process / hormone catabolic process / bradykinin catabolic process / cytosolic proteasome complex / insulin binding ...insulysin / beta-endorphin binding / ubiquitin recycling / insulin catabolic process / insulin metabolic process / amyloid-beta clearance by cellular catabolic process / hormone catabolic process / bradykinin catabolic process / cytosolic proteasome complex / insulin binding / regulation of aerobic respiration / negative regulation of glycogen catabolic process / positive regulation of nitric oxide mediated signal transduction / negative regulation of feeding behavior / negative regulation of fatty acid metabolic process / Signaling by Insulin receptor / peptide catabolic process / IRS activation / Insulin processing / regulation of protein secretion / positive regulation of peptide hormone secretion / positive regulation of respiratory burst / amyloid-beta clearance / Regulation of gene expression in beta cells / negative regulation of acute inflammatory response / peroxisomal matrix / alpha-beta T cell activation / positive regulation of dendritic spine maintenance / Synthesis, secretion, and deacylation of Ghrelin / negative regulation of respiratory burst involved in inflammatory response / amyloid-beta metabolic process / negative regulation of protein secretion / activation of protein kinase B activity / negative regulation of gluconeogenesis / positive regulation of insulin receptor signaling pathway / positive regulation of glycogen biosynthetic process / fatty acid homeostasis / Signal attenuation / positive regulation of protein binding / FOXO-mediated transcription of oxidative stress, metabolic and neuronal genes / negative regulation of lipid catabolic process / positive regulation of lipid biosynthetic process / regulation of protein localization to plasma membrane / negative regulation of oxidative stress-induced intrinsic apoptotic signaling pathway / transport vesicle / nitric oxide-cGMP-mediated signaling / COPI-mediated anterograde transport / positive regulation of nitric-oxide synthase activity / Insulin receptor recycling / negative regulation of reactive oxygen species biosynthetic process / insulin-like growth factor receptor binding / positive regulation of brown fat cell differentiation / NPAS4 regulates expression of target genes / negative regulation of proteolysis / endoplasmic reticulum-Golgi intermediate compartment membrane / neuron projection maintenance / peptide binding / positive regulation of mitotic nuclear division / Insulin receptor signalling cascade / proteolysis involved in protein catabolic process / Peroxisomal protein import / positive regulation of glycolytic process / positive regulation of cytokine production / positive regulation of long-term synaptic potentiation / endosome lumen / acute-phase response / positive regulation of protein secretion / positive regulation of D-glucose import / insulin receptor binding / positive regulation of cell differentiation / Regulation of insulin secretion / wound healing / protein catabolic process / negative regulation of protein catabolic process / antigen processing and presentation of endogenous peptide antigen via MHC class I / hormone activity / positive regulation of neuron projection development / regulation of synaptic plasticity / metalloendopeptidase activity / positive regulation of protein localization to nucleus / Golgi lumen / cognition / glucose metabolic process / positive regulation of protein catabolic process / vasodilation / peroxisome / insulin receptor signaling pathway / glucose homeostasis / cell-cell signaling / regulation of protein localization / amyloid-beta binding / virus receptor activity / PI5P, PP2A and IER3 Regulate PI3K/AKT Signaling / positive regulation of cell growth / protease binding / secretory granule lumen / endopeptidase activity / basolateral plasma membrane / positive regulation of canonical NF-kappaB signal transduction / Ub-specific processing proteases
Similarity search - Function
Peptidase M16, middle/third domain / Middle or third domain of peptidase_M16 / : / PQQ synthase PqqF-like, C-terminal lobe domain 4 / : / Peptidase M16, zinc-binding site / Insulinase family, zinc-binding region signature. / Peptidase M16, C-terminal / Peptidase M16 inactive domain / Peptidase M16, N-terminal ...Peptidase M16, middle/third domain / Middle or third domain of peptidase_M16 / : / PQQ synthase PqqF-like, C-terminal lobe domain 4 / : / Peptidase M16, zinc-binding site / Insulinase family, zinc-binding region signature. / Peptidase M16, C-terminal / Peptidase M16 inactive domain / Peptidase M16, N-terminal / Insulinase (Peptidase family M16) / Metalloenzyme, LuxS/M16 peptidase-like / Insulin / Insulin family / Insulin-like / Insulin/IGF/Relaxin family / Insulin / insulin-like growth factor / relaxin family. / Insulin, conserved site / Insulin family signature. / Insulin-like superfamily
Similarity search - Domain/homology
Insulin / Insulin-degrading enzyme
Similarity search - Component
Biological speciesHomo sapiens (human)
Methodsingle particle reconstruction / cryo EM / Resolution: 3.7 Å
AuthorsZhang Z / Liang WG / Bailey LJ / Tan YZ / Wei H / Kossiakoff AA / Carragher B / Potter SC / Tang WJ
Funding support United States, 4 items
OrganizationGrant numberCountry
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)GM103310 United States
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)GM81539 United States
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)GM121964 United States
Simons Foundation349247 United States
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- ...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.
History
DepositionSep 22, 2017-
Header (metadata) releaseNov 22, 2017-
Map releaseNov 22, 2017-
UpdateNov 13, 2024-
Current statusNov 13, 2024Processing site: RCSB / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 0.053
  • Imaged by UCSF Chimera
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  • Surface view colored by radius
  • Surface level: 0.053
  • Imaged by UCSF Chimera
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  • Surface view with fitted model
  • Atomic models: PDB-6b3q
  • Surface level: 0.053
  • Imaged by UCSF Chimera
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  • Surface view with fitted model
  • Atomic models: PDB-6bfc
  • Surface level: 0.053
  • Imaged by UCSF Chimera
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Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

emd_7041.png

Downloads & links

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Map

FileDownload / File: emd_7041.map.gz / Format: CCP4 / Size: 125 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationThis is the sharpened map from the final, Insulin degrading enzyme in complex with insulin
Projections & slices

Image control

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

Surface

Projections

Slices (1/3)

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Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 1.073 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.053 / Movie #1: 0.053
Minimum - Maximum-0.23219995 - 0.18164448
Average (Standard dev.)0.00027707088 (±0.0042306143)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions320320320
Spacing320320320
CellA=B=C: 343.36 Å
α=β=γ: 90.0 °

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
MAP C/R/S123
start NC/NR/NS000
NC/NR/NS320320320
D min/max/mean-0.2320.1820.000

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Supplemental data

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Mask #1

Fileemd_7041_msk_1.map
Projections & Slices
AxesZYX

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Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Additional map: This is the full, unfiltered map from final...

Fileemd_7041_additional.map
AnnotationThis is the full, unfiltered map from final refinement for insulin degrading enzyme in complex with insulin
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: This is the first half map from the...

Fileemd_7041_half_map_1.map
AnnotationThis is the first half map from the final refinement for insulin degrading enzyme in complex with insulin
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: This is the second half map from the...

Fileemd_7041_half_map_2.map
AnnotationThis is the second half map from the final refinement for insulin degrading enzyme in complex with insulin
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Sample components

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Entire : Insulin degrading enzyme/Insulin

EntireName: Insulin degrading enzyme/Insulin
Components
  • Complex: Insulin degrading enzyme/Insulin
    • Protein or peptide: Insulin-degrading enzyme
    • Protein or peptide: Insulin

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Supramolecule #1: Insulin degrading enzyme/Insulin

SupramoleculeName: Insulin degrading enzyme/Insulin / type: complex / ID: 1 / Parent: 0 / Macromolecule list: all
Details: Cryo-EM structure of human insulin degrading enzyme in complex with insulin
Source (natural)Organism: Homo sapiens (human)
Molecular weightTheoretical: 100 KDa

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Macromolecule #1: Insulin-degrading enzyme

MacromoleculeName: Insulin-degrading enzyme / type: protein_or_peptide / ID: 1 / Number of copies: 2 / Enantiomer: LEVO / EC number: insulysin
Source (natural)Organism: Homo sapiens (human)
Molecular weightTheoretical: 114.561562 KDa
Recombinant expressionOrganism: Escherichia coli (E. coli)
SequenceString: MHHHHHHAAG IPMNNPAIKR IGNHITKSPE DKREYRGLEL ANGIKVLLIS DPTTDKSSAA LDVHIGSLSD PPNIAGLSHF LEHMLFLGT KKYPKENEYS QFLSEHAGSS NAFTSGEHTN YYFDVSHEHL EGALDRFAQF FLSPLFDESA KDREVNAVDS E HEKNVMND ...String:
MHHHHHHAAG IPMNNPAIKR IGNHITKSPE DKREYRGLEL ANGIKVLLIS DPTTDKSSAA LDVHIGSLSD PPNIAGLSHF LEHMLFLGT KKYPKENEYS QFLSEHAGSS NAFTSGEHTN YYFDVSHEHL EGALDRFAQF FLSPLFDESA KDREVNAVDS E HEKNVMND AWRLFQLEKA TGNPKHPFSK FGTGNKYTLE TRPNQEGIDV RQELLKFHSA YYSSNLMAVV VLGRESLDDL TN LVVKLFS EVENKNVPLP EFPEHPFQEE HLKQLYKIVP IKDIRNLYVT FPIPDLQKYY KSNPGHYLGH LIGHEGPGSL LSE LKSKGW VNTLVGGQKE GARGFMFFII NVDLTEEGLL HVEDIILHMF QYIQKLRAEG PQEWVFQELK DLNAVAFRFK DKER PRGYT SKIAGILHYY PLEEVLTAEY LLEEFRPDLI EMVLDKLRPE NVRVAIVSKS FEGKTDRTEE WYGTQYKQEA IPDEV IKKW QNADLNGKFK LPTKNEFIPT NFEILPLEKE ATPYPALIKD TAMSKLWFKQ DDKFFLPKAN LNFEFFSPFA YVDPLH SNM AYLYLELLKD SLNEYAYAAE LAGLSYDLQN TIYGMYLSVK GYNDKQPILL KKIIEKMATF EIDEKRFEII KEAYMRS LN NFRAEQPHQH AMYYLRLLMT EVAWTKDELK EALDDVTLPR LKAFIPQLLS RLHIEALLHG NITKQAALGI MQMVEDTL I EHAHTKPLLP SQLVRYREVQ LPDRGWFVYQ QRNEVHNNSG IEIYYQTDMQ STSENMFLEL FAQIISEPAF NTLRTKEQL GYIVFSGPRR ANGIQGLRFI IQSEKPPHYL ESRVEAFLIT MEKSIEDMTE EAFQKHIQAL AIRRLDKPKK LSAESAKYWG EIISQQYNF DRDNTEVAYL KTLTKEDIIK FYKEMLAVDA PRRHKVSVHV LAREMDSNPV VGEFPAQNDI NLSQAPALPQ P EVIQNMTE FKRGLPLFPL VKPHINFMAA KL

UniProtKB: Insulin-degrading enzyme

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Macromolecule #2: Insulin

MacromoleculeName: Insulin / type: protein_or_peptide / ID: 2 / Number of copies: 2 / Enantiomer: LEVO
Source (natural)Organism: Homo sapiens (human)
Molecular weightTheoretical: 11.989862 KDa
Recombinant expressionOrganism: Escherichia coli (E. coli)
SequenceString:
MALWMRLLPL LALLALWGPD PAAAFVNQHL CGSHLVEALY LVCGERGFFY TPKTRREAED LQVGQVELGG GPGAGSLQPL ALEGSLQKR GIVEQCCTSI CSLYQLENYC N

UniProtKB: Insulin

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Experimental details

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Structure determination

Methodcryo EM
Processingsingle particle reconstruction
Aggregation stateparticle

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Sample preparation

Concentration0.3 mg/mL
BufferpH: 7.8
Component:
ConcentrationFormulaName
20.0 mmol/LC8H18N2O4SHEPES
300.0 mmol/LNaClSodium chloride
20.0 mmol/LC10H16N2O8EDTA
GridModel: homemade nanowire grid / Material: COPPER / Mesh: 300 / Support film - Material: CARBON / Support film - topology: HOLEY / Support film - Film thickness: 10 / Pretreatment - Type: GLOW DISCHARGE / Pretreatment - Time: 10 sec. / Pretreatment - Atmosphere: OTHER / Pretreatment - Pressure: 0.001 kPa
VitrificationCryogen name: ETHANE / Chamber humidity: 85 % / Chamber temperature: 298 K / Instrument: HOMEMADE PLUNGER
Details: The cryo grids were made using Spotiton and homemade plunger.
DetailsThe sample was monodisperse

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Electron microscopy

MicroscopeFEI TITAN KRIOS
TemperatureMin: 70.0 K / Max: 70.0 K
Alignment procedureComa free - Residual tilt: 10.0 mrad
Image recordingFilm or detector model: GATAN K2 SUMMIT (4k x 4k) / Detector mode: COUNTING / Digitization - Dimensions - Width: 3710 pixel / Digitization - Dimensions - Height: 3838 pixel / Digitization - Frames/image: 1-50 / Number grids imaged: 3 / Number real images: 3085 / Average exposure time: 10.0 sec. / Average electron dose: 71.4 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsC2 aperture diameter: 70.0 µm / Calibrated magnification: 46598 / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Cs: 2.7 mm / Nominal defocus max: 2.2 µm / Nominal defocus min: 0.9400000000000001 µm / Nominal magnification: 22500
Sample stageSpecimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Cooling holder cryogen: NITROGEN
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Particle selectionNumber selected: 762283
Startup modelType of model: PDB ENTRY
PDB model - PDB ID:

3qz2

Final reconstructionNumber classes used: 1 / Applied symmetry - Point group: C1 (asymmetric) / Algorithm: FOURIER SPACE / Resolution.type: BY AUTHOR / Resolution: 3.7 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: RELION (ver. 2.1) / Number images used: 116122
Initial angle assignmentType: PROJECTION MATCHING
Projection matching processing - Number reference projections: 2
Software - Name: RELION (ver. 2.0)
Final angle assignmentType: PROJECTION MATCHING
Projection matching processing - Number reference projections: 2
Software - Name: RELION (ver. 2.0)
Final 3D classificationNumber classes: 8 / Avg.num./class: 18549 / Software - Name: RELION (ver. 2.0)
FSC plot (resolution estimation)

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Atomic model buiding 1

RefinementSpace: REAL / Protocol: FLEXIBLE FIT / Overall B value: 92
Output model

PDB-6b3q:
Cryo-EM structure of human insulin degrading enzyme in complex with insulin

PDB-6bfc:
Cryo-EM structure of human insulin degrading enzyme in complex with insulin

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