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Basic information

Entry
Database: PDB / ID: 5wob
TitleCrystal Structure Analysis of Fab1-Bound Human Insulin Degrading Enzyme (IDE) in Complex with Insulin
Components
  • IDE-bound Fab heavy chain
  • IDE-bound Fab light chain
  • Insulin
  • Insulin-degrading enzyme
KeywordsHYDROLASE / 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 / negative regulation of NAD(P)H oxidase activity / regulation of aerobic respiration ...insulysin / ubiquitin recycling / insulin catabolic process / insulin metabolic process / amyloid-beta clearance by cellular catabolic process / hormone catabolic process / bradykinin catabolic process / insulin binding / negative regulation of NAD(P)H oxidase activity / regulation of aerobic respiration / negative regulation of glycogen catabolic process / positive regulation of nitric oxide mediated signal transduction / negative regulation of fatty acid metabolic process / Signaling by Insulin receptor / negative regulation of feeding behavior / 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 / positive regulation of protein autophosphorylation / 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 / positive regulation of glycogen biosynthetic process / negative regulation of gluconeogenesis / Signal attenuation / fatty acid homeostasis / FOXO-mediated transcription of oxidative stress, metabolic and neuronal genes / positive regulation of insulin receptor signaling pathway / negative regulation of lipid catabolic process / regulation of protein localization to plasma membrane / positive regulation of lipid biosynthetic process / negative regulation of oxidative stress-induced intrinsic apoptotic signaling pathway / activation of protein kinase B activity / COPI-mediated anterograde transport / transport vesicle / nitric oxide-cGMP-mediated signaling / negative regulation of reactive oxygen species biosynthetic process / Insulin receptor recycling / insulin-like growth factor receptor binding / positive regulation of brown fat cell differentiation / NPAS4 regulates expression of target genes / neuron projection maintenance / endoplasmic reticulum-Golgi intermediate compartment membrane / positive regulation of nitric-oxide synthase activity / positive regulation of mitotic nuclear division / Insulin receptor signalling cascade / regulation of transmembrane transporter activity / peptide binding / proteolysis involved in protein catabolic process / positive regulation of glycolytic process / positive regulation of long-term synaptic potentiation / endosome lumen / positive regulation of cytokine production / acute-phase response / positive regulation of protein secretion / positive regulation of D-glucose import / positive regulation of cell differentiation / Regulation of insulin secretion / insulin receptor binding / Peroxisomal protein import / 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 / regulation of synaptic plasticity / positive regulation of neuron projection development / metalloendopeptidase activity / positive regulation of protein localization to nucleus / Golgi lumen / cognition / glucose metabolic process / positive regulation of protein catabolic process / positive regulation of protein binding / vasodilation / peroxisome / insulin receptor signaling pathway / glucose homeostasis / cell-cell signaling / regulation of protein localization / virus receptor activity / PI5P, PP2A and IER3 Regulate PI3K/AKT Signaling / protease binding / positive regulation of cell growth / secretory granule lumen / basolateral plasma membrane / endopeptidase activity / positive regulation of canonical NF-kappaB signal transduction / positive regulation of phosphatidylinositol 3-kinase/protein kinase B signal transduction / positive regulation of MAPK cascade
Similarity search - Function
Peptidase M16, middle/third domain / Middle or third domain of peptidase_M16 / : / PQQ synthase PqqF-like, C-terminal lobe domain 4 / : / Cytochrome Bc1 Complex; Chain A, domain 1 / Metalloenzyme, LuxS/M16 peptidase-like / Peptidase M16, zinc-binding site / Insulinase family, zinc-binding region signature. / Peptidase M16, C-terminal ...Peptidase M16, middle/third domain / Middle or third domain of peptidase_M16 / : / PQQ synthase PqqF-like, C-terminal lobe domain 4 / : / Cytochrome Bc1 Complex; Chain A, domain 1 / Metalloenzyme, LuxS/M16 peptidase-like / 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 / Immunoglobulins / Immunoglobulin-like / Sandwich / 2-Layer Sandwich / Mainly Beta / Alpha Beta
Similarity search - Domain/homology
Insulin / Insulin-degrading enzyme
Similarity search - Component
Biological speciesHomo sapiens (human)
Mus musculoides (Temminck's mouse)
MethodX-RAY DIFFRACTION / SYNCHROTRON / Resolution: 3.95 Å
AuthorsMcCord, L.A. / Liang, W.G. / Farcasanu, M. / Wang, A.G. / Koide, S. / Tang, W.J.
Funding support United States, 1items
OrganizationGrant numberCountry
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)GM121964 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
DepositionAug 1, 2017Deposition site: RCSB / Processing site: RCSB
SupersessionApr 18, 2018ID: 4Q5Z
Revision 1.0Apr 18, 2018Provider: repository / Type: Initial release
Revision 1.1Oct 30, 2019Group: Data collection / Database references / Category: citation / citation_author
Item: _citation.country / _citation.journal_abbrev ..._citation.country / _citation.journal_abbrev / _citation.journal_id_CSD / _citation.journal_id_ISSN / _citation.journal_volume / _citation.pdbx_database_id_DOI / _citation.pdbx_database_id_PubMed / _citation.title / _citation.year
Revision 1.2Mar 23, 2022Group: Author supporting evidence / Database references / Category: database_2 / pdbx_audit_support
Item: _database_2.pdbx_DOI / _database_2.pdbx_database_accession ..._database_2.pdbx_DOI / _database_2.pdbx_database_accession / _pdbx_audit_support.funding_organization / _pdbx_audit_support.grant_number
Revision 1.3Oct 4, 2023Group: Data collection / Refinement description
Category: chem_comp_atom / chem_comp_bond / pdbx_initial_refinement_model
Revision 1.4Oct 23, 2024Group: Structure summary / Category: pdbx_entry_details / pdbx_modification_feature

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

Downloads & links

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Assembly

Deposited unit
A: Insulin-degrading enzyme
B: Insulin-degrading enzyme
C: Insulin-degrading enzyme
D: Insulin-degrading enzyme
E: Insulin-degrading enzyme
F: Insulin-degrading enzyme
G: Insulin-degrading enzyme
H: Insulin-degrading enzyme
a: Insulin
b: Insulin
c: Insulin
d: Insulin
e: Insulin
f: Insulin
g: Insulin
h: Insulin
I: IDE-bound Fab heavy chain
K: IDE-bound Fab heavy chain
M: IDE-bound Fab heavy chain
O: IDE-bound Fab heavy chain
Q: IDE-bound Fab heavy chain
S: IDE-bound Fab heavy chain
U: IDE-bound Fab heavy chain
W: IDE-bound Fab heavy chain
J: IDE-bound Fab light chain
L: IDE-bound Fab light chain
N: IDE-bound Fab light chain
P: IDE-bound Fab light chain
R: IDE-bound Fab light chain
T: IDE-bound Fab light chain
V: IDE-bound Fab light chain
X: IDE-bound Fab light chain
hetero molecules


Theoretical massNumber of molelcules
Total (without water)1,368,63540
Polymers1,368,11232
Non-polymers5238
Water00
1
A: Insulin-degrading enzyme
B: Insulin-degrading enzyme
a: Insulin
b: Insulin
I: IDE-bound Fab heavy chain
K: IDE-bound Fab heavy chain
J: IDE-bound Fab light chain
L: IDE-bound Fab light chain
hetero molecules


Theoretical massNumber of molelcules
Total (without water)342,15910
Polymers342,0288
Non-polymers1312
Water0
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1
2
C: Insulin-degrading enzyme
D: Insulin-degrading enzyme
c: Insulin
d: Insulin
M: IDE-bound Fab heavy chain
O: IDE-bound Fab heavy chain
N: IDE-bound Fab light chain
P: IDE-bound Fab light chain
hetero molecules


Theoretical massNumber of molelcules
Total (without water)342,15910
Polymers342,0288
Non-polymers1312
Water0
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1
3
E: Insulin-degrading enzyme
F: Insulin-degrading enzyme
e: Insulin
f: Insulin
Q: IDE-bound Fab heavy chain
S: IDE-bound Fab heavy chain
R: IDE-bound Fab light chain
T: IDE-bound Fab light chain
hetero molecules


Theoretical massNumber of molelcules
Total (without water)342,15910
Polymers342,0288
Non-polymers1312
Water0
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1
4
G: Insulin-degrading enzyme
H: Insulin-degrading enzyme
g: Insulin
h: Insulin
U: IDE-bound Fab heavy chain
W: IDE-bound Fab heavy chain
V: IDE-bound Fab light chain
X: IDE-bound Fab light chain
hetero molecules


Theoretical massNumber of molelcules
Total (without water)342,15910
Polymers342,0288
Non-polymers1312
Water0
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1
Unit cell
Length a, b, c (Å)121.589, 138.190, 376.509
Angle α, β, γ (deg.)90.00, 99.36, 90.00
Int Tables number4
Space group name H-MP1211

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Components

#1: Protein
Insulin-degrading enzyme / Abeta-degrading protease / Insulin protease / Insulinase / Insulysin


Mass: 114560.578 Da / Num. of mol.: 8 / Fragment: UNP residues 42-1019
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: IDE / Production host: Escherichia coli (E. coli) / References: UniProt: P14735, insulysin
#2: Protein/peptide
Insulin


Mass: 2269.595 Da / Num. of mol.: 8
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: INS / Production host: Saccharomyces cerevisiae (brewer's yeast) / References: UniProt: P01308
#3: Antibody
IDE-bound Fab heavy chain


Mass: 28201.670 Da / Num. of mol.: 8
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Mus musculoides (Temminck's mouse) / Production host: Escherichia coli (E. coli)
#4: Antibody
IDE-bound Fab light chain


Mass: 25982.098 Da / Num. of mol.: 8
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Mus musculoides (Temminck's mouse) / Production host: Escherichia coli (E. coli)
#5: Chemical
ChemComp-ZN / ZINC ION


Mass: 65.409 Da / Num. of mol.: 8 / Source method: obtained synthetically / Formula: Zn / Feature type: SUBJECT OF INVESTIGATION
Has protein modificationY

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Experimental details

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Experiment

ExperimentMethod: X-RAY DIFFRACTION / Number of used crystals: 1

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Sample preparation

CrystalDensity Matthews: 2.28 Å3/Da / Density % sol: 46.08 %
Crystal growTemperature: 291.15 K / Method: vapor diffusion, hanging drop / pH: 6.5
Details: 0.1M Sodium cacodylate, pH6.5; 0.2M MgCl2; 10% PEG3000, VAPOR DIFFUSION, HANGING DROP, temperature 291.15K

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Data collection

DiffractionMean temperature: 100 K
Diffraction sourceSource: SYNCHROTRON / Site: APS / Beamline: 19-ID / Wavelength: 0.9792 Å
DetectorType: ADSC QUANTUM 315r / Detector: CCD / Date: Jul 11, 2013
RadiationProtocol: SINGLE WAVELENGTH / Monochromatic (M) / Laue (L): M / Scattering type: x-ray
Radiation wavelengthWavelength: 0.9792 Å / Relative weight: 1
ReflectionResolution: 3.95→50 Å / Num. obs: 108370 / % possible obs: 99.5 % / Observed criterion σ(F): 2.1 / Observed criterion σ(I): 2.1 / Redundancy: 3.3 % / Rmerge(I) obs: 0.2 / Rpim(I) all: 0.13 / Rsym value: 0.12 / Χ2: 1.277 / Net I/σ(I): 7
Reflection shellResolution: 3.95→4.02 Å / Redundancy: 3.3 % / Rmerge(I) obs: 0.672 / Mean I/σ(I) obs: 2.1 / Num. unique obs: 5406 / CC1/2: 0.583 / Rpim(I) all: 0.424 / Rsym value: 0.621 / Χ2: 1.02 / % possible all: 99.8

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Processing

Software
NameVersionClassification
PHENIX1.10.1_2155refinement
HKL-2000data reduction
HKL-2000data scaling
PHASERphasing
RefinementStarting model: 4IOF

4iof


Resolution: 3.95→49.543 Å / Cross valid method: FREE R-VALUE / σ(F): 1.34 / Phase error: 26.49
RfactorNum. reflection% reflection
Rfree0.291 1994 1.85 %
Rwork0.2431 --
obs0.2439 107673 99.45 %
Solvent computationShrinkage radii: 0.9 Å / VDW probe radii: 1.11 Å
Refinement stepCycle: LAST / Resolution: 3.95→49.543 Å
ProteinNucleic acidLigandSolventTotal
Num. atoms86898 0 8 0 86906
Refine LS restraints
Refine-IDTypeDev idealNumber
X-RAY DIFFRACTIONf_bond_d0.00288973
X-RAY DIFFRACTIONf_angle_d0.453120368
X-RAY DIFFRACTIONf_dihedral_angle_d15.42153581
X-RAY DIFFRACTIONf_chiral_restr0.03913128
X-RAY DIFFRACTIONf_plane_restr0.00415391
LS refinement shell
Resolution (Å)Rfactor RfreeNum. reflection RfreeRfactor RworkNum. reflection RworkRefine-ID% reflection obs (%)
3.9505-4.04930.30941450.28367569X-RAY DIFFRACTION98
4.0493-4.15870.31041420.27427464X-RAY DIFFRACTION98
4.1587-4.2810.33811410.26437555X-RAY DIFFRACTION98
4.281-4.41910.34871380.25757487X-RAY DIFFRACTION98
4.4191-4.57690.28871420.24037578X-RAY DIFFRACTION98
4.5769-4.760.26081410.22797537X-RAY DIFFRACTION98
4.76-4.97650.25531420.21227536X-RAY DIFFRACTION98
4.9765-5.23860.25911390.22257537X-RAY DIFFRACTION98
5.2386-5.56640.27911420.24067589X-RAY DIFFRACTION98
5.5664-5.99550.34431390.25627546X-RAY DIFFRACTION98
5.9955-6.59760.31741410.28237570X-RAY DIFFRACTION98
6.5976-7.54940.29091420.25147541X-RAY DIFFRACTION97
7.5494-9.50040.2521400.20457462X-RAY DIFFRACTION96
9.5004-49.54670.29751430.24127694X-RAY DIFFRACTION97

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