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- EMDB-13388: Low resolution Cryo-EM structure of the full-length insulin recep... -

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Basic information

Entry
Database: EMDB / ID: EMD-13388
TitleLow resolution Cryo-EM structure of the full-length insulin receptor bound to 2 insulin, conf 3
Map data
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 / exocrine pancreas development / insulin receptor complex / insulin-like growth factor I binding / insulin receptor activity / positive regulation of protein-containing complex disassembly ...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 / exocrine pancreas development / insulin receptor complex / insulin-like growth factor I binding / insulin receptor activity / positive regulation of protein-containing complex disassembly / cargo receptor activity / dendritic spine maintenance / insulin binding / negative regulation of NAD(P)H oxidase activity / negative regulation of glycogen catabolic process / PTB domain binding / adrenal gland development / positive regulation of nitric oxide mediated signal transduction / negative regulation of fatty acid metabolic process / activation of protein kinase activity / negative regulation of feeding behavior / Signaling by Insulin receptor / IRS activation / Insulin processing / neuronal cell body membrane / regulation of protein secretion / positive regulation of peptide hormone secretion / positive regulation of respiratory burst / positive regulation of receptor internalization / negative regulation of acute inflammatory response / Regulation of gene expression in beta cells / alpha-beta T cell activation / amyloid-beta clearance / regulation of amino acid metabolic process / regulation of embryonic development / negative regulation of respiratory burst involved in inflammatory response / insulin receptor substrate binding / negative regulation of protein secretion / positive regulation of dendritic spine maintenance / transport across blood-brain barrier / positive regulation of glycogen biosynthetic process / Synthesis, secretion, and deacylation of Ghrelin / epidermis development / regulation of protein localization to plasma membrane / fatty acid homeostasis / negative regulation of lipid catabolic process / negative regulation of gluconeogenesis / Signal attenuation / FOXO-mediated transcription of oxidative stress, metabolic and neuronal genes / COPI-mediated anterograde transport / phosphatidylinositol 3-kinase binding / positive regulation of lipid biosynthetic process / heart morphogenesis / negative regulation of oxidative stress-induced intrinsic apoptotic signaling pathway / positive regulation of insulin receptor signaling pathway / nitric oxide-cGMP-mediated signaling / negative regulation of reactive oxygen species biosynthetic process / positive regulation of protein autophosphorylation / transport vesicle / Insulin receptor recycling / insulin-like growth factor receptor binding / dendrite membrane / neuron projection maintenance / endoplasmic reticulum-Golgi intermediate compartment membrane / positive regulation of brown fat cell differentiation / positive regulation of protein metabolic process / NPAS4 regulates expression of target genes / activation of protein kinase B activity / positive regulation of glycolytic process / positive regulation of mitotic nuclear division / Insulin receptor signalling cascade / receptor-mediated endocytosis / positive regulation of nitric-oxide synthase activity / learning / positive regulation of cytokine production / positive regulation of long-term synaptic potentiation / acute-phase response / endosome lumen / Regulation of insulin secretion / positive regulation of D-glucose import / positive regulation of protein secretion / negative regulation of proteolysis / positive regulation of cell differentiation / regulation of transmembrane transporter activity / insulin receptor binding / positive regulation of MAP kinase activity / wound healing / receptor protein-tyrosine kinase / caveola / regulation of synaptic plasticity / negative regulation of protein catabolic process / cellular response to growth factor stimulus / hormone activity / receptor internalization / memory / positive regulation of neuron projection development / peptidyl-tyrosine phosphorylation / cellular response to insulin stimulus / cognition / positive regulation of protein localization to nucleus
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: 9.1 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-
UpdateNov 6, 2024-
Current statusNov 6, 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-7pg4
  • Surface level: 0.125
  • Imaged by UCSF Chimera
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Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

emd_13388.png

Downloads & links

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Map

FileDownload / File: emd_13388.map.gz / Format: CCP4 / Size: 282.6 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
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-0.09111873 - 0.39998895
Average (Standard dev.)0.0013307504 (±0.0154407285)
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-0.0910.4000.001

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Supplemental data

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Mask #1

Fileemd_13388_msk_1.map
Projections & Slices
AxesZYX

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Slices (1/2)
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Histogram

Histogram (log scale)

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Half map: #1

Fileemd_13388_half_map_1.map
Projections & Slices
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Density Histograms

Histogram

Histogram (log scale)

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Half map: #2

Fileemd_13388_half_map_2.map
Projections & Slices
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Histogram (log scale)

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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
Source (natural)Organism: Homo sapiens (human)
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: 2 / 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: 2 / 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: 9.1 Å / Resolution method: OTHER / Software - Name: cryoSPARC (ver. 2.15) / Details: Masked FSC calculated with GSFSC in cryoSPARC2. / Number images used: 26701
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-7pg4:
Low resolution Cryo-EM structure of the full-length insulin receptor bound to 2 insulin, conf 3

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