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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: EMD-7066
TitleCryo-EM structure of human insulin degrading enzyme in complex with insulin
Map dataInsulin degrading enzyme in complex with insulin
Sample
  • Complex: Insulin degrading enzyme
    • Protein or peptide: Insulin-degrading enzyme
    • Protein or peptide: FAB H11 heavy chain
    • Protein or peptide: FAB H11 light chain
KeywordsIDE / insulin degrading enzyme / amyloid beta / BIOSYNTHETIC PROTEIN / HYDROLASE-IMMUNE SYSTEM complex
Function / homology
Function and homology information


insulysin / ubiquitin recycling / insulin catabolic process / insulin metabolic process / amyloid-beta clearance by cellular catabolic process / hormone catabolic process / bradykinin catabolic process / insulin binding / regulation of aerobic respiration / peptide catabolic process ...insulysin / ubiquitin recycling / insulin catabolic process / insulin metabolic process / amyloid-beta clearance by cellular catabolic process / hormone catabolic process / bradykinin catabolic process / insulin binding / regulation of aerobic respiration / peptide catabolic process / immunoglobulin complex / amyloid-beta clearance / peroxisomal matrix / amyloid-beta metabolic process / Insulin receptor recycling / proteolysis involved in protein catabolic process / Peroxisomal protein import / peptide binding / protein catabolic process / antigen processing and presentation of endogenous peptide antigen via MHC class I / metalloendopeptidase activity / positive regulation of protein catabolic process / positive regulation of protein binding / peroxisome / insulin receptor signaling pathway / virus receptor activity / basolateral plasma membrane / endopeptidase activity / adaptive immune response / Ub-specific processing proteases / immune response / external side of plasma membrane / cell surface / protein homodimerization activity / mitochondrion / proteolysis / extracellular space / extracellular exosome / zinc ion binding / extracellular region / ATP binding / identical protein binding / nucleus / plasma membrane / cytosol / cytoplasm
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 / Insulinase (Peptidase family M16) ...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 / : / : / Immunoglobulin V-Type / Immunoglobulin V-set domain / Immunoglobulin V-set domain / Immunoglobulin subtype / Immunoglobulin / Immunoglobulin/major histocompatibility complex, conserved site / Immunoglobulins and major histocompatibility complex proteins signature. / Immunoglobulin C-Type / Immunoglobulin C1-set / Immunoglobulin C1-set domain / Ig-like domain profile. / Immunoglobulin-like domain / Immunoglobulin-like domain superfamily / Immunoglobulin-like fold
Similarity search - Domain/homology
Immunoglobulin gamma-1 heavy chain / Insulin-degrading enzyme / Ig-like domain-containing protein
Similarity search - Component
Biological speciesHomo sapiens (human)
Methodsingle particle reconstruction / cryo EM / Resolution: 6.5 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
DepositionOct 5, 2017-
Header (metadata) releaseJan 10, 2018-
Map releaseJan 10, 2018-
UpdateOct 9, 2024-
Current statusOct 9, 2024Processing site: RCSB / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 0.04
  • Imaged by UCSF Chimera
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  • 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

emd_7066.png

Downloads & links

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Map

FileDownload / File: emd_7066.map.gz / Format: CCP4 / Size: 125 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationInsulin 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)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 1.073 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.04 / Movie #1: 0.04
Minimum - Maximum-0.27929044 - 0.35023555
Average (Standard dev.)0.00038742728 (±0.007850708)
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-0.2790.3500.000

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Supplemental data

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Mask #1

Fileemd_7066_msk_1.map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: Insulin degrading enzyme in complex with insulin

Fileemd_7066_half_map_1.map
AnnotationInsulin 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: Insulin degrading enzyme in complex with insulin

Fileemd_7066_half_map_2.map
AnnotationInsulin 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

EntireName: Insulin degrading enzyme
Components
  • Complex: Insulin degrading enzyme
    • Protein or peptide: Insulin-degrading enzyme
    • Protein or peptide: FAB H11 heavy chain
    • Protein or peptide: FAB H11 light chain

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Supramolecule #1: Insulin degrading enzyme

SupramoleculeName: Insulin degrading enzyme / type: complex / ID: 1 / Parent: 0 / Macromolecule list: all
Details: Cryo-EM structure of human Apo insulin degrading enzyme
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: 111.866484 KDa
Recombinant expressionOrganism: Escherichia coli (E. coli)
SequenceString: AIKRIGNHIT KSPEDKREYR GLELANGIKV LLISDPTTDK SSAALDVHIG SLSDPPNIAG LSHFLEHMLF LGTKKYPKEN EYSQFLSEH AGSSNAFTSG EHTNYYFDVS HEHLEGALDR FAQFFLSPLF DESAKDREVN AVDSEHEKNV MNDAWRLFQL E KATGNPKH ...String:
AIKRIGNHIT KSPEDKREYR GLELANGIKV LLISDPTTDK SSAALDVHIG SLSDPPNIAG LSHFLEHMLF LGTKKYPKEN EYSQFLSEH AGSSNAFTSG EHTNYYFDVS HEHLEGALDR FAQFFLSPLF DESAKDREVN AVDSEHEKNV MNDAWRLFQL E KATGNPKH PFSKFGTGNK YTLETRPNQE GIDVRQELLK FHSAYYSSNL MAVVVLGRES LDDLTNLVVK LFSEVENKNV PL PEFPEHP FQEEHLKQLY KIVPIKDIRN LYVTFPIPDL QKYYKSNPGH YLGHLIGHEG PGSLLSELKS KGWVNTLVGG QKE GARGFM FFIINVDLTE EGLLHVEDII LHMFQYIQKL RAEGPQEWVF QELKDLNAVA FRFKDKERPR GYTSKIAGIL HYYP LEEVL TAEYLLEEFR PDLIEMVLDK LRPENVRVAI VSKSFEGKTD RTEEWYGTQY KQEAIPDEVI KKWQNADLNG KFKLP TKNE FIPTNFEILP LEKEATPYPA LIKDTAMSKL WFKQDDKFFL PKANLNFEFF SPFAYVDPLH SNMAYLYLEL LKDSLN EYA YAAELAGLSY DLQNTIYGMY LSVKGYNDKQ PILLKKIIEK MATFEIDEKR FEIIKEAYMR SLNNFRAEQP HQHAMYY LR LLMTEVAWTK DELKEALDDV TLPRLKAFIP QLLSRLHIEA LLHGNITKQA ALGIMQMVED TLIEHAHTKP LLPSQLVR Y REVQLPDRGW FVYQQRNEVH NNSGIEIYYQ TDMQSTSENM FLELFAQIIS EPAFNTLRTK EQLGYIVFSG PRRANGIQG LRFIIQSEKP PHYLESRVEA FLITMEKSIE DMTEEAFQKH IQALAIRRLD KPKKLSAESA KYWGEIISQQ YNFDRDNTEV AYLKTLTKE DIIKFYKEML AVDAPRRHKV SVHVLAREMD SCPVVGEFPC QNDINLSQAP ALPQPEVIQN MTEFKRGLPL F PLVKPH

UniProtKB: Insulin-degrading enzyme

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Macromolecule #2: FAB H11 heavy chain

MacromoleculeName: FAB H11 heavy chain / type: protein_or_peptide / ID: 2 / Number of copies: 2 / Enantiomer: LEVO
Source (natural)Organism: Homo sapiens (human)
Molecular weightTheoretical: 23.057748 KDa
Recombinant expressionOrganism: Escherichia coli (E. coli)
SequenceString: EVQLVESGGG LVQPGGSLRL SCAASGFNIS SSSIHWVRQA PGKGLEWVAS IYSYSGSTYY ADSVKGRFTI SADTSKNTAY LQMNSLRAE DTAVYYCARH YSAVAGLDYW GQGTLVTVFN QIKPPSVFPL APSSKSTSGG TAALGCLVKD YFPEPVTVSW N SGALTSGV ...String:
EVQLVESGGG LVQPGGSLRL SCAASGFNIS SSSIHWVRQA PGKGLEWVAS IYSYSGSTYY ADSVKGRFTI SADTSKNTAY LQMNSLRAE DTAVYYCARH YSAVAGLDYW GQGTLVTVFN QIKPPSVFPL APSSKSTSGG TAALGCLVKD YFPEPVTVSW N SGALTSGV HTFPAVLQSS GLYSLSSVVT VPSSSLGTQT YICNVNHKPS NTKVDKKVEP

UniProtKB: Immunoglobulin gamma-1 heavy chain

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Macromolecule #3: FAB H11 light chain

MacromoleculeName: FAB H11 light chain / type: protein_or_peptide / ID: 3 / Number of copies: 2 / Enantiomer: LEVO
Source (natural)Organism: Homo sapiens (human)
Molecular weightTheoretical: 23.087609 KDa
Recombinant expressionOrganism: Escherichia coli (E. coli)
SequenceString: DIQMTQSPSS LSASVGDRVT ITCRASQSVS SAVAWYQQKP GKAPKLLIYS ASSLYSGVPS RFSGSRSGTD YTLTISSLQP EDFATYYCQ QSYFNPITFG QGTKVEIKRT VAAPSVFIFP PSDEQLKSGT ASVVCLLNNF YPREAKVQWK VDNALQSGNS Q ESVTEQDS ...String:
DIQMTQSPSS LSASVGDRVT ITCRASQSVS SAVAWYQQKP GKAPKLLIYS ASSLYSGVPS RFSGSRSGTD YTLTISSLQP EDFATYYCQ QSYFNPITFG QGTKVEIKRT VAAPSVFIFP PSDEQLKSGT ASVVCLLNNF YPREAKVQWK VDNALQSGNS Q ESVTEQDS KDSTYSLSST LTLSKADYEK HKVYACEVTH QGLSSPVTKS FNR

UniProtKB: Ig-like domain-containing protein

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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 / Material: GOLD / 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 / Details: The grids are homemade lacey gold nanowire grids
VitrificationCryogen name: ETHANE / Chamber humidity: 85 % / Chamber temperature: 298 K / Instrument: HOMEMADE PLUNGER / Details: The cryo grids were made using Spotiton.
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
DetailsThe image was collected at 20-50 degree tilt
Image recording#0 - Image recording ID: 1 / #0 - Film or detector model: GATAN K2 SUMMIT (4k x 4k) / #0 - Detector mode: COUNTING / #0 - Number grids imaged: 1 / #0 - Number real images: 509 / #0 - Average exposure time: 10.0 sec. / #0 - Average electron dose: 7.9 e/Å2 / #1 - Image recording ID: 2 / #1 - Film or detector model: GATAN K2 SUMMIT (4k x 4k) / #1 - Detector mode: COUNTING / #1 - Digitization - Dimensions - Width: 3710 pixel / #1 - Digitization - Dimensions - Height: 3820 pixel / #1 - Digitization - Frames/image: 1-50 / #1 - Number grids imaged: 1 / #1 - Number real images: 620 / #1 - Average exposure time: 10.0 sec. / #1 - Average electron dose: 6.8 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

Image recording ID1
Particle selectionNumber selected: 762283
Startup modelType of model: PDB ENTRY
PDB model - PDB ID:

4q5z
PDB Unreleased entry

Final reconstructionNumber classes used: 1 / Algorithm: FOURIER SPACE / Resolution.type: BY AUTHOR / Resolution: 6.5 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: RELION (ver. 2.1) / Number images used: 16944
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: 18983 / 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-6b7z:
Cryo-EM structure of human insulin degrading enzyme in complex with FAB H11 heavy chain and FAB H11 light chain

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