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- EMDB-13385: Low resolution Cryo-EM structure of full-length insulin receptor ... -

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Basic information

Entry
Database: EMDB / ID: EMD-13385
TitleLow resolution Cryo-EM structure of full-length insulin receptor bound to 3 insulin with visible ddm micelle, conf 1
Map dataDDM solubilised full-length human insulin receptor with three insulins bound with visible detergent micelle.
Sample
  • Complex: DDM solubilised full-length human insulin receptor with three insulins bound
    • Protein or peptide: Isoform Short of Insulin receptor
    • Protein or peptide: Insulin
    • Protein or peptide: Insulin
KeywordsInsulin / Receptor / Complex / MEMBRANE PROTEIN
Function / homology
Function and homology information


regulation of female gonad development / positive regulation of meiotic cell cycle / insulin-like growth factor II binding / positive regulation of developmental growth / male sex determination / insulin receptor complex / insulin-like growth factor I binding / exocrine pancreas development / positive regulation of protein-containing complex disassembly / dendritic spine maintenance ...regulation of female gonad development / positive regulation of meiotic cell cycle / insulin-like growth factor II binding / positive regulation of developmental growth / male sex determination / insulin receptor complex / insulin-like growth factor I binding / exocrine pancreas development / positive regulation of protein-containing complex disassembly / dendritic spine maintenance / cargo receptor activity / insulin binding / negative regulation of NAD(P)H oxidase activity / neuronal cell body membrane / adrenal gland development / PTB domain binding / 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 / 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 / positive regulation of protein autophosphorylation / regulation of embryonic development / alpha-beta T cell activation / positive regulation of receptor internalization / protein kinase activator activity / insulin receptor substrate binding / positive regulation of dendritic spine maintenance / Synthesis, secretion, and deacylation of Ghrelin / negative regulation of respiratory burst involved in inflammatory response / epidermis development / negative regulation of protein secretion / negative regulation of gluconeogenesis / positive regulation of glycogen biosynthetic process / Signal attenuation / fatty acid homeostasis / phosphatidylinositol 3-kinase binding / FOXO-mediated transcription of oxidative stress, metabolic and neuronal genes / positive regulation of insulin receptor signaling pathway / negative regulation of lipid catabolic process / transport across blood-brain barrier / regulation of protein localization to plasma membrane / positive regulation of lipid biosynthetic process / heart morphogenesis / 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 / dendrite membrane / NPAS4 regulates expression of target genes / neuron projection maintenance / endoplasmic reticulum-Golgi intermediate compartment membrane / positive regulation of MAP kinase activity / positive regulation of nitric-oxide synthase activity / positive regulation of mitotic nuclear division / Insulin receptor signalling cascade / receptor-mediated endocytosis / regulation of transmembrane transporter activity / positive regulation of glycolytic process / learning / positive regulation of long-term synaptic potentiation / positive regulation of cytokine production / endosome lumen / acute-phase response / positive regulation of D-glucose import / positive regulation of protein secretion / positive regulation of cell differentiation / Regulation of insulin secretion / insulin receptor binding / wound healing / placental growth factor receptor activity / insulin receptor activity / vascular endothelial growth factor receptor activity / hepatocyte growth factor receptor activity / macrophage colony-stimulating factor receptor activity / platelet-derived growth factor alpha-receptor activity / platelet-derived growth factor beta-receptor activity / stem cell factor receptor activity / boss receptor activity / protein tyrosine kinase collagen receptor activity / brain-derived neurotrophic factor receptor activity / transmembrane-ephrin receptor activity / GPI-linked ephrin receptor activity / epidermal growth factor receptor activity / fibroblast growth factor receptor activity / insulin-like growth factor receptor activity / receptor protein-tyrosine kinase
Similarity search - Function
Insulin receptor, trans-membrane domain / Insulin receptor trans-membrane segment / Tyrosine-protein kinase, insulin-like receptor / Tyrosine-protein kinase, receptor class II, conserved site / Receptor tyrosine kinase class II signature. / Insulin / Insulin family / Insulin-like / Insulin/IGF/Relaxin family / Insulin / insulin-like growth factor / relaxin family. ...Insulin receptor, trans-membrane domain / Insulin receptor trans-membrane segment / Tyrosine-protein kinase, insulin-like receptor / Tyrosine-protein kinase, receptor class II, conserved site / Receptor tyrosine kinase class II signature. / 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 / Receptor L-domain / Furin-like cysteine-rich domain / Receptor L-domain superfamily / Furin-like cysteine rich region / Receptor L domain / Furin-like repeat / Furin-like repeats / Growth factor receptor cysteine-rich domain superfamily / Fibronectin type III domain / : / Fibronectin type 3 domain / Fibronectin type-III domain profile. / Fibronectin type III / Fibronectin type III superfamily / Tyrosine-protein kinase, catalytic domain / Tyrosine kinase, catalytic domain / Tyrosine protein kinases specific active-site signature. / Tyrosine-protein kinase, active site / Serine-threonine/tyrosine-protein kinase, catalytic domain / Protein tyrosine and serine/threonine kinase / Protein kinase, ATP binding site / Protein kinases ATP-binding region signature. / Immunoglobulin-like fold / Protein kinase domain profile. / Protein kinase domain / Protein kinase-like domain superfamily
Similarity search - Domain/homology
Insulin / Insulin receptor
Similarity search - Component
Biological speciesHomo sapiens (human)
Methodsingle particle reconstruction / cryo EM / Resolution: 7.6 Å
AuthorsNielsen JA / Slaaby R
Funding support Denmark, 1 items
OrganizationGrant numberCountry
Other government Denmark
CitationJournal: J Mol Biol / Year: 2022
Title: Structural Investigations of Full-Length Insulin Receptor Dynamics and Signalling.
Authors: Jeppe Nielsen / Jakob Brandt / Thomas Boesen / Tina Hummelshøj / Rita Slaaby / Gerd Schluckebier / Poul Nissen /
Abstract: Insulin regulates glucose homeostasis via binding and activation of the insulin receptor dimer at two distinct pairs of binding sites 1 and 2. Here, we present cryo-EM studies of full-length human ...Insulin regulates glucose homeostasis via binding and activation of the insulin receptor dimer at two distinct pairs of binding sites 1 and 2. Here, we present cryo-EM studies of full-length human insulin receptor (hIR) in an active state obtained at non-saturating, physiologically relevant insulin conditions. Insulin binds asymmetrically to the receptor under these conditions, occupying up to three of the four possible binding sites. Deletion analysis of the receptor together with site specific peptides and insulin analogs used in binding studies show that both sites 1 and 2 are required for high insulin affinity. We identify a homotypic interaction of the fibronectin type III domain (FnIII-3) of IR resulting in tight interaction of membrane proximal domains of the active, asymmetric receptor dimer. Our results show how insulin binding at two distinct types of sites disrupts the autoinhibited apo-IR dimer and stabilizes the active dimer. We propose an insulin binding and activation mechanism, which is sequential, exhibits negative cooperativity, and is based on asymmetry at physiological insulin concentrations with one to three insulin molecules activating IR.
History
DepositionAug 12, 2021-
Header (metadata) releaseFeb 2, 2022-
Map releaseFeb 2, 2022-
UpdateOct 23, 2024-
Current statusOct 23, 2024Processing site: PDBe / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 0.125
  • Imaged by UCSF Chimera
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  • Surface view colored by cylindrical radius
  • Surface level: 0.125
  • Imaged by UCSF Chimera
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  • Surface view with fitted model
  • Atomic models: PDB-7pg0
  • Surface level: 0.125
  • Imaged by UCSF Chimera
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  • Simplified surface model + fitted atomic model
  • Atomic modelsPDB-7pg0
  • Imaged by Jmol
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Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

emd_13385.png

Downloads & links

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Map

FileDownload / File: emd_13385.map.gz / Format: CCP4 / Size: 282.6 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationDDM solubilised full-length human insulin receptor with three insulins bound with visible detergent micelle.
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
1.09 Å/pix.
x 420 pix.
= 457.716 Å		1.09 Å/pix.
x 420 pix.
= 457.716 Å		1.09 Å/pix.
x 420 pix.
= 457.716 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 1.0898 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.125 / Movie #1: 0.125
Minimum - Maximum-0.037651476 - 0.31327534
Average (Standard dev.)0.001349606 (±0.013307925)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions420420420
Spacing420420420
CellA=B=C: 457.716 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z1.08981.08981.0898
M x/y/z420420420
origin x/y/z0.0000.0000.000
length x/y/z457.716457.716457.716
α/β/γ90.00090.00090.000
MAP C/R/S123
start NC/NR/NS000
NC/NR/NS420420420
D min/max/mean-0.0380.3130.001

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Supplemental data

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Mask #1

Fileemd_13385_msk_1.map
Projections & Slices
AxesZYX

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Density Histograms

Histogram

Histogram (log scale)

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Half map: #2

Fileemd_13385_half_map_1.map
Projections & Slices
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Slices (1/2)
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Histogram

Histogram (log scale)

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Half map: #1

Fileemd_13385_half_map_2.map
Projections & Slices
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Sample components

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Entire : DDM solubilised full-length human insulin receptor with three ins...

EntireName: DDM solubilised full-length human insulin receptor with three insulins bound
Components
  • Complex: DDM solubilised full-length human insulin receptor with three insulins bound
    • Protein or peptide: Isoform Short of Insulin receptor
    • Protein or peptide: Insulin
    • Protein or peptide: Insulin

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Supramolecule #1: DDM solubilised full-length human insulin receptor with three ins...

SupramoleculeName: DDM solubilised full-length human insulin receptor with three insulins bound
type: complex / ID: 1 / Parent: 0 / Macromolecule list: all
Molecular weightTheoretical: 460 KDa

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Macromolecule #1: Isoform Short of Insulin receptor

MacromoleculeName: Isoform Short of Insulin receptor / type: protein_or_peptide / ID: 1 / Number of copies: 2 / Enantiomer: LEVO / EC number: receptor protein-tyrosine kinase
Source (natural)Organism: Homo sapiens (human)
Molecular weightTheoretical: 156.697578 KDa
Recombinant expressionOrganism: Cricetulus griseus (Chinese hamster)
SequenceString: MATGGRRGAA AAPLLVAVAA LLLGAAGHLY PGEVCPGMDI RNNLTRLHEL ENCSVIEGHL QILLMFKTRP EDFRDLSFPK LIMITDYLL LFRVYGLESL KDLFPNLTVI RGSRLFFNYA LVIFEMVHLK ELGLYNLMNI TRGSVRIEKN NELCYLATID W SRILDSVE ...String:
MATGGRRGAA AAPLLVAVAA LLLGAAGHLY PGEVCPGMDI RNNLTRLHEL ENCSVIEGHL QILLMFKTRP EDFRDLSFPK LIMITDYLL LFRVYGLESL KDLFPNLTVI RGSRLFFNYA LVIFEMVHLK ELGLYNLMNI TRGSVRIEKN NELCYLATID W SRILDSVE DNYIVLNKDD NEECGDICPG TAKGKTNCPA TVINGQFVER CWTHSHCQKV CPTICKSHGC TAEGLCCHSE CL GNCSQPD DPTKCVACRN FYLDGRCVET CPPPYYHFQD WRCVNFSFCQ DLHHKCKNSR RQGCHQYVIH NNKCIPECPS GYT MNSSNL LCTPCLGPCP KVCHLLEGEK TIDSVTSAQE LRGCTVINGS LIINIRGGNN LAAELEANLG LIEEISGYLK IRRS YALVS LSFFRKLRLI RGETLEIGNY SFYALDNQNL RQLWDWSKHN LTITQGKLFF HYNPKLCLSE IHKMEEVSGT KGRQE RNDI ALKTNGDQAS CENELLKFSY IRTSFDKILL RWEPYWPPDF RDLLGFMLFY KEAPYQNVTE FDGQDACGSN SWTVVD IDP PLRSNDPKSQ NHPGWLMRGL KPWTQYAIFV KTLVTFSDER RTYGAKSDII YVQTDATNPS VPLDPISVSN SSSQIIL KW KPPSDPNGNI THYLVFWERQ AEDSELFELD YCLKGLKLPS RTWSPPFESE DSQKHNQSEY EDSAGECCSC PKTDSQIL K ELEESSFRKT FEDYLHNVVF VPRPSRKRRS LGDVGNVTVA VPTVAAFPNT SSTSVPTSPE EHRPFEKVVN KESLVISGL RHFTGYRIEL QACNQDTPEE RCSVAAYVSA RTMPEAKADD IVGPVTHEIF ENNVVHLMWQ EPKEPNGLIV LYEVSYRRYG DEELHLCVS RKHFALERGC RLRGLSPGNY SVRIRATSLA GNGSWTEPTY FYVTDYLDVP SNIAKIIIGP LIFVFLFSVV I GSIYLFLR KRQPDGPLGP LYASSNPEYL SASDVFPCSV YVPDEWEVSR EKITLLRELG QGSFGMVYEG NARDIIKGEA ET RVAVKTV NESASLRERI EFLNEASVMK GFTCHHVVRL LGVVSKGQPT LVVMELMAHG DLKSYLRSLR PEAENNPGRP PPT LQEMIQ MAAEIADGMA YLNAKKFVHR DLAARNCMVA HDFTVKIGDF GMTRDIYETD YYRKGGKGLL PVRWMAPESL KDGV FTTSS DMWSFGVVLW EITSLAEQPY QGLSNEQVLK FVMDGGYLDQ PDNCPERVTD LMRMCWQFNP KMRPTFLEIV NLLKD DLHP SFPEVSFFHS EENKAPESEE LEMEFEDMEN VPLDRSSHCQ REEAGGRDGG SSLGFKRSYE EHIPYTHMNG GKKNGR ILT LPRSNPSEDQ VDPRLIDGK

UniProtKB: Insulin receptor

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Macromolecule #2: Insulin

MacromoleculeName: Insulin / type: protein_or_peptide / ID: 2 / Number of copies: 3 / Enantiomer: LEVO
Source (natural)Organism: Homo sapiens (human)
Molecular weightTheoretical: 2.383698 KDa
Recombinant expressionOrganism: Saccharomyces cerevisiae (brewer's yeast)
SequenceString:
GIVEQCCTSI CSLYQLENYC N

UniProtKB: Insulin

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Macromolecule #3: Insulin

MacromoleculeName: Insulin / type: protein_or_peptide / ID: 3 / Number of copies: 3 / Enantiomer: LEVO
Source (natural)Organism: Homo sapiens (human)
Molecular weightTheoretical: 3.433953 KDa
Recombinant expressionOrganism: Saccharomyces cerevisiae (brewer's yeast)
SequenceString:
FVNQHLCGSH LVEALYLVCG ERGFFYTPKT

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

Concentration4.0 mg/mL
BufferpH: 7.8
Component:
ConcentrationFormulaName
150.0 mMNaClSodium Chloride
50.0 mMC8H18N2O4SHepes
0.03 w/v%C24H46O11DDM
GridModel: C-flat-2/2 / Material: COPPER / Support film - Material: CARBON / Support film - topology: HOLEY ARRAY / Pretreatment - Type: GLOW DISCHARGE
VitrificationCryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 297 K / Instrument: LEICA EM GP / Details: Blotted for 3s prior to plunging.

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Image recordingFilm or detector model: GATAN K2 SUMMIT (4k x 4k) / Average exposure time: 15.0 sec. / Average electron dose: 60.0 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Nominal defocus max: 2.0 µm / Nominal defocus min: 0.5 µm
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Startup modelType of model: NONE
Final reconstructionResolution.type: BY AUTHOR / Resolution: 7.6 Å / Resolution method: OTHER / Software - Name: cryoSPARC (ver. 2.15) / Details: Masked FSC calculated with GSFSC in cryoSPARC2. / Number images used: 32763
Initial angle assignmentType: MAXIMUM LIKELIHOOD
Final angle assignmentType: MAXIMUM LIKELIHOOD
FSC plot (resolution estimation)

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Atomic model buiding 1

RefinementProtocol: FLEXIBLE FIT
Output model

PDB-7pg0:
Low resolution Cryo-EM structure of full-length insulin receptor bound to 3 insulin with visible ddm micelle, conf 1

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