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- EMDB-8978: Cryo-EM structure of the active, Gs-protein complexed, human CGRP... -

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Basic information

Entry
Database: EMDB / ID: EMD-8978
TitleCryo-EM structure of the active, Gs-protein complexed, human CGRP receptor
Map dataActive, Gs-protein complexed, CGRP receptor
Sample
  • Complex: Complex of a full-length, active-state calcitonin gene-related peptide receptor with calcitonin gene-related peptide ligand, heterotrimeric Gs protein and nanobody35
    • Protein or peptide: Calcitonin gene-related peptide 1
    • Protein or peptide: Nanobody 35Single-domain antibody
    • Protein or peptide: Guanine nucleotide-binding protein G(s) subunit alpha isoforms short
    • Protein or peptide: Guanine nucleotide-binding protein G(I)/G(S)/G(T) subunit beta-1
    • Protein or peptide: Guanine nucleotide-binding protein G(I)/G(S)/G(O) subunit gamma-2
    • Protein or peptide: Calcitonin gene-related peptide type 1 receptor
    • Protein or peptide: Receptor activity-modifying protein 1
Function / homology
Function and homology information


nervous system process involved in regulation of systemic arterial blood pressure / calcitonin gene-related peptide binding / CGRP receptor complex / calcitonin gene-related peptide receptor signaling pathway / adrenomedullin binding / positive regulation of protein glycosylation / cellular response to sucrose stimulus / adrenomedullin receptor activity / adrenomedullin receptor complex / adrenomedullin receptor signaling pathway ...nervous system process involved in regulation of systemic arterial blood pressure / calcitonin gene-related peptide binding / CGRP receptor complex / calcitonin gene-related peptide receptor signaling pathway / adrenomedullin binding / positive regulation of protein glycosylation / cellular response to sucrose stimulus / adrenomedullin receptor activity / adrenomedullin receptor complex / adrenomedullin receptor signaling pathway / amylin receptor activity / calcitonin receptor activity / positive regulation of interleukin-1 alpha production / vascular associated smooth muscle cell proliferation / calcitonin gene-related peptide receptor activity / amylin receptor signaling pathway / Calcitonin-like ligand receptors / negative regulation of calcium ion transport into cytosol / positive regulation of macrophage differentiation / regulation of G protein-coupled receptor signaling pathway / G protein-coupled receptor internalization / vasculature development / endothelial cell proliferation / negative regulation of bone resorption / leukocyte cell-cell adhesion / negative regulation of osteoclast differentiation / PKA activation in glucagon signalling / G protein-coupled receptor signaling pathway, coupled to cyclic nucleotide second messenger / hair follicle placode formation / developmental growth / D1 dopamine receptor binding / endothelial cell migration / intracellular transport / Hedgehog 'off' state / positive regulation of cAMP-mediated signaling / coreceptor activity / adenylate cyclase-activating adrenergic receptor signaling pathway / cellular response to hormone stimulus / positive regulation of vascular associated smooth muscle cell proliferation / regulation of cytosolic calcium ion concentration / activation of adenylate cyclase activity / adenylate cyclase activator activity / negative regulation of blood pressure / trans-Golgi network membrane / protein localization to plasma membrane / G protein-coupled receptor activity / positive regulation of interleukin-8 production / intracellular protein transport / Olfactory Signaling Pathway / G-protein beta/gamma-subunit complex binding / Activation of the phototransduction cascade / bone development / G beta:gamma signalling through PLC beta / Presynaptic function of Kainate receptors / Thromboxane signalling through TP receptor / G-protein activation / G protein-coupled acetylcholine receptor signaling pathway / Activation of G protein gated Potassium channels / Inhibition of voltage gated Ca2+ channels via Gbeta/gamma subunits / adenylate cyclase-activating G protein-coupled receptor signaling pathway / Prostacyclin signalling through prostacyclin receptor / Glucagon signaling in metabolic regulation / G beta:gamma signalling through CDC42 / receptor internalization / hormone activity / ADP signalling through P2Y purinoceptor 12 / G beta:gamma signalling through BTK / Synthesis, secretion, and inactivation of Glucagon-like Peptide-1 (GLP-1) / Sensory perception of sweet, bitter, and umami (glutamate) taste / photoreceptor disc membrane / cognition / platelet aggregation / Adrenaline,noradrenaline inhibits insulin secretion / Glucagon-type ligand receptors / positive regulation of GTPase activity / regulation of blood pressure / Vasopressin regulates renal water homeostasis via Aquaporins / G alpha (z) signalling events / cellular response to catecholamine stimulus / Glucagon-like Peptide-1 (GLP1) regulates insulin secretion / ADORA2B mediated anti-inflammatory cytokines production / ADP signalling through P2Y purinoceptor 1 / adenylate cyclase-activating dopamine receptor signaling pathway / G beta:gamma signalling through PI3Kgamma / cellular response to prostaglandin E stimulus / Cooperation of PDCL (PhLP1) and TRiC/CCT in G-protein beta folding / vasodilation / sensory perception of taste / GPER1 signaling / G-protein beta-subunit binding / Inactivation, recovery and regulation of the phototransduction cascade / calcium ion transport / heterotrimeric G-protein complex / G alpha (12/13) signalling events / extracellular vesicle / sensory perception of smell / signaling receptor complex adaptor activity / Thrombin signalling through proteinase activated receptors (PARs) / protein transport / cell-cell signaling
Similarity search - Function
Calcitonin gene-related peptide / GPCR, family 2, calcitonin gene-related peptide, type 1 receptor / Receptor activity modifying protein / RAMP domain superfamily / Receptor activity modifying family / GPCR, family 2, calcitonin receptor family / Calcitonin-like / Calcitonin peptide-like / Calcitonin, conserved site / Calcitonin / CGRP / IAPP family signature. ...Calcitonin gene-related peptide / GPCR, family 2, calcitonin gene-related peptide, type 1 receptor / Receptor activity modifying protein / RAMP domain superfamily / Receptor activity modifying family / GPCR, family 2, calcitonin receptor family / Calcitonin-like / Calcitonin peptide-like / Calcitonin, conserved site / Calcitonin / CGRP / IAPP family signature. / calcitonin / Calcitonin/adrenomedullin / Calcitonin / CGRP / IAPP family / G-protein coupled receptors family 2 signature 1. / Hormone receptor domain / GPCR, family 2, extracellular hormone receptor domain / G-protein coupled receptors family 2 profile 1. / Domain present in hormone receptors / GPCR family 2, extracellular hormone receptor domain superfamily / G-protein coupled receptors family 2 signature 2. / GPCR, family 2, secretin-like, conserved site / GPCR, family 2, secretin-like / 7 transmembrane receptor (Secretin family) / GPCR, family 2-like / G-protein coupled receptors family 2 profile 2. / G-protein alpha subunit, group S / G-alpha domain profile. / Guanine nucleotide binding protein (G-protein), alpha subunit / G protein alpha subunit, helical insertion / G-protein alpha subunit / G protein alpha subunit / G-protein, gamma subunit / G-protein gamma subunit domain profile. / GGL domain / G-protein gamma-like domain superfamily / G-protein gamma-like domain / GGL domain / G protein gamma subunit-like motifs / Guanine nucleotide-binding protein, beta subunit / G-protein, beta subunit / G-protein beta WD-40 repeat / WD40 repeat, conserved site / Trp-Asp (WD) repeats signature. / WD domain, G-beta repeat / WD40 repeats / WD40 repeat / Trp-Asp (WD) repeats profile. / Trp-Asp (WD) repeats circular profile. / WD40-repeat-containing domain superfamily / WD40/YVTN repeat-like-containing domain superfamily / P-loop containing nucleoside triphosphate hydrolase
Similarity search - Domain/homology
Receptor activity-modifying protein 1 / Calcitonin gene-related peptide 1 / Guanine nucleotide-binding protein G(I)/G(S)/G(O) subunit gamma-2 / Guanine nucleotide-binding protein G(I)/G(S)/G(T) subunit beta-1 / Guanine nucleotide-binding protein G(s) subunit alpha isoforms short / Calcitonin gene-related peptide type 1 receptor
Similarity search - Component
Biological speciesHomo sapiens (human) / Lama glama (llama)
Methodsingle particle reconstruction / cryo EM / Resolution: 3.3 Å
AuthorsLiang YL / Khoshouei M / Deganutti G / Glukhova A / Koole C / Peat TS / Radjainia M / Plitzko JM / Baumeister W / Miller LJ ...Liang YL / Khoshouei M / Deganutti G / Glukhova A / Koole C / Peat TS / Radjainia M / Plitzko JM / Baumeister W / Miller LJ / Hay DL / Christopoulos A / Reynolds CA / Wootten D / Sexton PM
Funding support Australia, 2 items
OrganizationGrant numberCountry
National Health and Medical Research Council (NHMRC, Australia)1120919 Australia
National Health and Medical Research Council (NHMRC, Australia)1055134 Australia
CitationJournal: Nature / Year: 2018
Title: Cryo-EM structure of the active, G-protein complexed, human CGRP receptor.
Authors: Yi-Lynn Liang / Maryam Khoshouei / Giuseppe Deganutti / Alisa Glukhova / Cassandra Koole / Thomas S Peat / Mazdak Radjainia / Jürgen M Plitzko / Wolfgang Baumeister / Laurence J Miller / ...Authors: Yi-Lynn Liang / Maryam Khoshouei / Giuseppe Deganutti / Alisa Glukhova / Cassandra Koole / Thomas S Peat / Mazdak Radjainia / Jürgen M Plitzko / Wolfgang Baumeister / Laurence J Miller / Deborah L Hay / Arthur Christopoulos / Christopher A Reynolds / Denise Wootten / Patrick M Sexton /
Abstract: Calcitonin gene-related peptide (CGRP) is a widely expressed neuropeptide that has a major role in sensory neurotransmission. The CGRP receptor is a heterodimer of the calcitonin receptor-like ...Calcitonin gene-related peptide (CGRP) is a widely expressed neuropeptide that has a major role in sensory neurotransmission. The CGRP receptor is a heterodimer of the calcitonin receptor-like receptor (CLR) class B G-protein-coupled receptor and a type 1 transmembrane domain protein, receptor activity-modifying protein 1 (RAMP1). Here we report the structure of the human CGRP receptor in complex with CGRP and the G-protein heterotrimer at 3.3 Å global resolution, determined by Volta phase-plate cryo-electron microscopy. The receptor activity-modifying protein transmembrane domain sits at the interface between transmembrane domains 3, 4 and 5 of CLR, and stabilizes CLR extracellular loop 2. RAMP1 makes only limited direct contact with CGRP, consistent with its function in allosteric modulation of CLR. Molecular dynamics simulations indicate that RAMP1 provides stability to the receptor complex, particularly in the positioning of the extracellular domain of CLR. This work provides insights into the control of G-protein-coupled receptor function.
History
DepositionJul 16, 2018-
Header (metadata) releaseAug 1, 2018-
Map releaseSep 19, 2018-
UpdateJan 15, 2020-
Current statusJan 15, 2020Processing 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 height
  • Surface level: 0.04
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  • Surface view with fitted model
  • Atomic models: PDB-6e3y
  • Surface level: 0.04
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Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

Downloads & links

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Map

FileDownload / File: emd_8978.map.gz / Format: CCP4 / Size: 22.2 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationActive, Gs-protein complexed, CGRP receptor
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
1.06 Å/pix.
x 180 pix.
= 190.8 Å
1.06 Å/pix.
x 180 pix.
= 190.8 Å
1.06 Å/pix.
x 180 pix.
= 190.8 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 1.06 Å
Density
Contour LevelBy AUTHOR: 0.04 / Movie #1: 0.04
Minimum - Maximum-0.071706764 - 0.17341213
Average (Standard dev.)0.0015997638 (±0.008368622)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions180180180
Spacing180180180
CellA=B=C: 190.79999 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z1.061.061.06
M x/y/z180180180
origin x/y/z0.0000.0000.000
length x/y/z190.800190.800190.800
α/β/γ90.00090.00090.000
MAP C/R/S123
start NC/NR/NS000
NC/NR/NS180180180
D min/max/mean-0.0720.1730.002

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Supplemental data

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Sample components

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Entire : Complex of a full-length, active-state calcitonin gene-related pe...

EntireName: Complex of a full-length, active-state calcitonin gene-related peptide receptor with calcitonin gene-related peptide ligand, heterotrimeric Gs protein and nanobody35
Components
  • Complex: Complex of a full-length, active-state calcitonin gene-related peptide receptor with calcitonin gene-related peptide ligand, heterotrimeric Gs protein and nanobody35
    • Protein or peptide: Calcitonin gene-related peptide 1
    • Protein or peptide: Nanobody 35Single-domain antibody
    • Protein or peptide: Guanine nucleotide-binding protein G(s) subunit alpha isoforms short
    • Protein or peptide: Guanine nucleotide-binding protein G(I)/G(S)/G(T) subunit beta-1
    • Protein or peptide: Guanine nucleotide-binding protein G(I)/G(S)/G(O) subunit gamma-2
    • Protein or peptide: Calcitonin gene-related peptide type 1 receptor
    • Protein or peptide: Receptor activity-modifying protein 1

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Supramolecule #1: Complex of a full-length, active-state calcitonin gene-related pe...

SupramoleculeName: Complex of a full-length, active-state calcitonin gene-related peptide receptor with calcitonin gene-related peptide ligand, heterotrimeric Gs protein and nanobody35
type: complex / ID: 1 / Parent: 0 / Macromolecule list: all
Source (natural)Organism: Homo sapiens (human)
Recombinant expressionOrganism: Trichoplusia ni (cabbage looper)

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Macromolecule #1: Calcitonin gene-related peptide 1

MacromoleculeName: Calcitonin gene-related peptide 1 / type: protein_or_peptide / ID: 1 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Homo sapiens (human)
Molecular weightTheoretical: 3.795354 KDa
SequenceString:
ACDTATCVTH RLAGLLSRSG GVVKNNFVPT NVGSKAF(NH2)

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Macromolecule #2: Nanobody 35

MacromoleculeName: Nanobody 35 / type: protein_or_peptide / ID: 2 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Lama glama (llama)
Molecular weightTheoretical: 15.140742 KDa
Recombinant expressionOrganism: Escherichia coli (E. coli)
SequenceString:
QVQLQESGGG LVQPGGSLRL SCAASGFTFS NYKMNWVRQA PGKGLEWVSD ISQSGASISY TGSVKGRFTI SRDNAKNTLY LQMNSLKPE DTAVYYCARC PAPFTRDCFD VTSTTYAYRG QGTQVTVSSH HHHHHEPEA

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Macromolecule #3: Guanine nucleotide-binding protein G(s) subunit alpha isoforms short

MacromoleculeName: Guanine nucleotide-binding protein G(s) subunit alpha isoforms short
type: protein_or_peptide / ID: 3 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Homo sapiens (human)
Molecular weightTheoretical: 45.683434 KDa
Recombinant expressionOrganism: Trichoplusia ni (cabbage looper)
SequenceString: MGCLGNSKTE DQRNEEKAQR EANKKIEKQL QKDKQVYRAT HRLLLLGAGE SGKNTIVKQM RILHVNGFNG EGGEEDPQAA RSNSDGEKA TKVQDIKNNL KEAIETIVAA MSNLVPPVEL ANPENQFRVD YILSVMNVPD FDFPPEFYEH AKALWEDEGV R ACYERSNE ...String:
MGCLGNSKTE DQRNEEKAQR EANKKIEKQL QKDKQVYRAT HRLLLLGAGE SGKNTIVKQM RILHVNGFNG EGGEEDPQAA RSNSDGEKA TKVQDIKNNL KEAIETIVAA MSNLVPPVEL ANPENQFRVD YILSVMNVPD FDFPPEFYEH AKALWEDEGV R ACYERSNE YQLIDCAQYF LDKIDVIKQA DYVPSDQDLL RCRVLTSGIF ETKFQVDKVN FHMFDVGAQR DERRKWIQCF ND VTAIIFV VASSSYNMVI REDNQTNRLQ AALKLFDSIW NNKWLRDTSV ILFLNKQDLL AEKVLAGKSK IEDYFPEFAR YTT PEDATP EPGEDPRVTR AKYFIRDEFL RISTASGDGR HYCYPHFTCA VDTENIRRVF NDCRDIIQRM HLRQYELL

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Macromolecule #4: Guanine nucleotide-binding protein G(I)/G(S)/G(T) subunit beta-1

MacromoleculeName: Guanine nucleotide-binding protein G(I)/G(S)/G(T) subunit beta-1
type: protein_or_peptide / ID: 4 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Homo sapiens (human)
Molecular weightTheoretical: 38.534062 KDa
Recombinant expressionOrganism: Trichoplusia ni (cabbage looper)
SequenceString: MHHHHHHGSS GSELDQLRQE AEQLKNQIRD ARKACADATL SQITNNIDPV GRIQMRTRRT LRGHLAKIYA MHWGTDSRLL VSASQDGKL IIWDSYTTNK VHAIPLRSSW VMTCAYAPSG NYVACGGLDN ICSIYNLKTR EGNVRVSREL AGHTGYLSCC R FLDDNQIV ...String:
MHHHHHHGSS GSELDQLRQE AEQLKNQIRD ARKACADATL SQITNNIDPV GRIQMRTRRT LRGHLAKIYA MHWGTDSRLL VSASQDGKL IIWDSYTTNK VHAIPLRSSW VMTCAYAPSG NYVACGGLDN ICSIYNLKTR EGNVRVSREL AGHTGYLSCC R FLDDNQIV TSSGDTTCAL WDIETGQQTT TFTGHTGDVM SLSLAPDTRL FVSGACDASA KLWDVREGMC RQTFTGHESD IN AICFFPN GNAFATGSDD ATCRLFDLRA DQELMTYSHD NIICGITSVS FSKSGRLLLA GYDDFNCNVW DALKADRAGV LAG HDNRVS CLGVTDDGMA VATGSWDSFL KIWN

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Macromolecule #5: Guanine nucleotide-binding protein G(I)/G(S)/G(O) subunit gamma-2

MacromoleculeName: Guanine nucleotide-binding protein G(I)/G(S)/G(O) subunit gamma-2
type: protein_or_peptide / ID: 5 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Homo sapiens (human)
Molecular weightTheoretical: 7.861143 KDa
Recombinant expressionOrganism: Trichoplusia ni (cabbage looper)
SequenceString:
MASNNTASIA QARKLVEQLK MEANIDRIKV SKAAADLMAY CEAHAKEDPL LTPVPASENP FREKKFFCAI L

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Macromolecule #6: Calcitonin gene-related peptide type 1 receptor

MacromoleculeName: Calcitonin gene-related peptide type 1 receptor / type: protein_or_peptide / ID: 6 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Homo sapiens (human)
Molecular weightTheoretical: 56.27452 KDa
Recombinant expressionOrganism: Trichoplusia ni (cabbage looper)
SequenceString: MKTIIALSYI FCLVFADYKD DDDLEVLFQG PAELEESPED SIQLGVTRNK IMTAQYECYQ KIMQDPIQQA EGVYCNRTWD GWLCWNDVA AGTESMQLCP DYFQDFDPSE KVTKICDQDG NWFRHPASNR TWTNYTQCNV NTHEKVKTAL NLFYLTIIGH G LSIASLLI ...String:
MKTIIALSYI FCLVFADYKD DDDLEVLFQG PAELEESPED SIQLGVTRNK IMTAQYECYQ KIMQDPIQQA EGVYCNRTWD GWLCWNDVA AGTESMQLCP DYFQDFDPSE KVTKICDQDG NWFRHPASNR TWTNYTQCNV NTHEKVKTAL NLFYLTIIGH G LSIASLLI SLGIFFYFKS LSCQRITLHK NLFFSFVCNS VVTIIHLTAV ANNQALVATN PVSCKVSQFI HLYLMGCNYF WM LCEGIYL HTLIVVAVFA EKQHLMWYYF LGWGFPLIPA CIHAIARSLY YNDNCWISSD THLLYIIHGP ICAALLVNLF FLL NIVRVL ITKLKVTHQA ESNLYMKAVR ATLILVPLLG IEFVLIPWRP EGKIAEEVYD YIMHILMHFQ GLLVSTIFCF FNGE VQAIL RRNWNQYKIQ FGNSFSNSEA LRSASYTVST ISDGPGYSHD CPSEHLNGKS IHDIENVLLK PENLYNPAGL EVLFQ GPHH HHHHHH

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Macromolecule #7: Receptor activity-modifying protein 1

MacromoleculeName: Receptor activity-modifying protein 1 / type: protein_or_peptide / ID: 7 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Homo sapiens (human)
Molecular weightTheoretical: 17.066701 KDa
Recombinant expressionOrganism: Trichoplusia ni (cabbage looper)
SequenceString:
MKTIIALSYI FCLVFADYKD DDDKHGSCQE ANYGALLREL CLTQFQVDME AVGETLWCDW GRTIRSYREL ADCTWHMAEK LGCFWPNAE VDRFFLAVHG RYFRSCPISG RAVRDPPGSI LYPFIVVPIT VTLLVTALVV WQSKRTEGIV

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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

BufferpH: 7.4
GridModel: Quantifoil R1.2/1.3 / Material: COPPER / Mesh: 200
VitrificationCryogen name: NITROGEN / Chamber humidity: 100 % / Chamber temperature: 277 K / Instrument: FEI VITROBOT MARK IV

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsCalibrated magnification: 47170 / Illumination mode: OTHER / Imaging mode: BRIGHT FIELDBright-field microscopy
Sample stageSpecimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Cooling holder cryogen: NITROGEN
Image recordingFilm or detector model: GATAN K2 SUMMIT (4k x 4k) / Detector mode: COUNTING / Average electron dose: 50.0 e/Å2
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

CTF correctionDetails: phase plate CTF correction
Initial angle assignmentType: COMMON LINE
Final angle assignmentType: PROJECTION MATCHING
Final reconstructionApplied symmetry - Point group: C1 (asymmetric) / Resolution.type: BY AUTHOR / Resolution: 3.3 Å / Resolution method: FSC 0.143 CUT-OFF / Number images used: 407000

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