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- EMDB-25403: 5-HT2B receptor bound to LSD in complex with beta-arrestin1 obtai... -

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Basic information

Entry
Database: EMDB / ID: EMD-25403
Title5-HT2B receptor bound to LSD in complex with beta-arrestin1 obtained by cryo-electron microscopy (cryoEM)
Map dataComposite cryo-EM map for 5-HT2BR bound to LSD in complex with beta-arrestin1
Sample
  • Complex: 5-HT2B receptor bound to LSD in complex with beta-arrestin1
    • Protein or peptide: Anti-5HT2BR Fab light chain
    • Protein or peptide: Anti-5HT2BR Fab heavy chain
    • Protein or peptide: 5-hydroxytryptamine receptor 2B
    • Protein or peptide: Isoform 1B of Beta-arrestin-1
    • Protein or peptide: Fab30 heavy chain
    • Protein or peptide: Fab30 light chain
  • Ligand: 2-acetamido-2-deoxy-beta-D-glucopyranose
  • Ligand: (8alpha)-N,N-diethyl-6-methyl-9,10-didehydroergoline-8-carboxamide
Function / homology
Function and homology information


angiotensin receptor binding / Activation of SMO / negative regulation of interleukin-8 production / arrestin family protein binding / G protein-coupled receptor internalization / enzyme inhibitor activity / Lysosome Vesicle Biogenesis / positive regulation of Rho protein signal transduction / Golgi Associated Vesicle Biogenesis / negative regulation of NF-kappaB transcription factor activity ...angiotensin receptor binding / Activation of SMO / negative regulation of interleukin-8 production / arrestin family protein binding / G protein-coupled receptor internalization / enzyme inhibitor activity / Lysosome Vesicle Biogenesis / positive regulation of Rho protein signal transduction / Golgi Associated Vesicle Biogenesis / negative regulation of NF-kappaB transcription factor activity / stress fiber assembly / negative regulation of Notch signaling pathway / pseudopodium / negative regulation of interleukin-6 production / positive regulation of receptor internalization / clathrin-coated pit / negative regulation of protein ubiquitination / insulin-like growth factor receptor binding / visual perception / Activated NOTCH1 Transmits Signal to the Nucleus / GTPase activator activity / G protein-coupled receptor binding / Signaling by high-kinase activity BRAF mutants / MAP2K and MAPK activation / cytoplasmic vesicle membrane / Signaling by RAF1 mutants / Signaling by moderate kinase activity BRAF mutants / Paradoxical activation of RAF signaling by kinase inactive BRAF / Signaling downstream of RAS mutants / Signaling by BRAF and RAF1 fusions / Thrombin signalling through proteinase activated receptors (PARs) / Cargo recognition for clathrin-mediated endocytosis / protein transport / Clathrin-mediated endocytosis / ubiquitin-dependent protein catabolic process / cytoplasmic vesicle / G alpha (s) signalling events / proteasome-mediated ubiquitin-dependent protein catabolic process / transcription coactivator activity / positive regulation of ERK1 and ERK2 cascade / protein ubiquitination / nuclear body / Ub-specific processing proteases / positive regulation of protein phosphorylation / lysosomal membrane / Golgi membrane / ubiquitin protein ligase binding / chromatin / regulation of transcription by RNA polymerase II / signal transduction / positive regulation of transcription by RNA polymerase II / nucleoplasm / nucleus / plasma membrane / cytosol / cytoplasm
Similarity search - Function
Arrestin, conserved site / Arrestins signature. / Arrestin / Arrestin, N-terminal / Arrestin-like, N-terminal / Arrestin C-terminal-like domain / Arrestin (or S-antigen), N-terminal domain / Arrestin (or S-antigen), C-terminal domain / Arrestin (or S-antigen), C-terminal domain / Arrestin-like, C-terminal / Immunoglobulin E-set
Similarity search - Domain/homology
Biological speciesMus musculus (house mouse) / Homo sapiens (human)
Methodsingle particle reconstruction / cryo EM / Resolution: 3.3 Å
AuthorsBarros-Alvarez X / Cao C / Panova O / Roth BL / Skiniotis G
Funding support1 items
OrganizationGrant numberCountry
Other government
CitationJournal: Neuron / Year: 2022
Title: Signaling snapshots of a serotonin receptor activated by the prototypical psychedelic LSD.
Authors: Can Cao / Ximena Barros-Álvarez / Shicheng Zhang / Kuglae Kim / Marc A Dämgen / Ouliana Panova / Carl-Mikael Suomivuori / Jonathan F Fay / Xiaofang Zhong / Brian E Krumm / Ryan H Gumpper / ...Authors: Can Cao / Ximena Barros-Álvarez / Shicheng Zhang / Kuglae Kim / Marc A Dämgen / Ouliana Panova / Carl-Mikael Suomivuori / Jonathan F Fay / Xiaofang Zhong / Brian E Krumm / Ryan H Gumpper / Alpay B Seven / Michael J Robertson / Nevan J Krogan / Ruth Hüttenhain / David E Nichols / Ron O Dror / Georgios Skiniotis / Bryan L Roth /
Abstract: Serotonin (5-hydroxytryptamine [5-HT]) 5-HT2-family receptors represent essential targets for lysergic acid diethylamide (LSD) and all other psychedelic drugs. Although the primary psychedelic drug ...Serotonin (5-hydroxytryptamine [5-HT]) 5-HT2-family receptors represent essential targets for lysergic acid diethylamide (LSD) and all other psychedelic drugs. Although the primary psychedelic drug effects are mediated by the 5-HT serotonin receptor (HTR2A), the 5-HT serotonin receptor (HTR2B) has been used as a model receptor to study the activation mechanisms of psychedelic drugs due to its high expression and similarity to HTR2A. In this study, we determined the cryo-EM structures of LSD-bound HTR2B in the transducer-free, Gq-protein-coupled, and β-arrestin-1-coupled states. These structures provide distinct signaling snapshots of LSD's action, ranging from the transducer-free, partially active state to the transducer-coupled, fully active states. Insights from this study will both provide comprehensive molecular insights into the signaling mechanisms of the prototypical psychedelic LSD and accelerate the discovery of novel psychedelic drugs.
History
DepositionNov 8, 2021-
Header (metadata) releaseSep 21, 2022-
Map releaseSep 21, 2022-
UpdateOct 19, 2022-
Current statusOct 19, 2022Processing site: RCSB / Status: Released

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Structure visualization

Supplemental images

emd_25403.png

Downloads & links

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Map

FileDownload / File: emd_25403.map.gz / Format: CCP4 / Size: 178 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationComposite cryo-EM map for 5-HT2BR bound to LSD in complex with beta-arrestin1
Voxel sizeX=Y=Z: 0.8521 Å
Density
Contour LevelBy AUTHOR: 0.12
Minimum - Maximum-0.20221582 - 0.7823746
Average (Standard dev.)0.0007283417 (±0.011113655)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions360360360
Spacing360360360
CellA=B=C: 306.756 Å
α=β=γ: 90.0 °

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Supplemental data

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Additional map: Focused refinement map for 5-HT2BR bound to LSD...

Fileemd_25403_additional_1.map
AnnotationFocused refinement map for 5-HT2BR bound to LSD and a scFv fragment of anti-5HT2BR Fab
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Additional map: Focused refinement map for beta-arrestin1, scFv30 and 5HT2BR C-tail

Fileemd_25403_additional_2.map
AnnotationFocused refinement map for beta-arrestin1, scFv30 and 5HT2BR C-tail
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Sample components

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Entire : 5-HT2B receptor bound to LSD in complex with beta-arrestin1

EntireName: 5-HT2B receptor bound to LSD in complex with beta-arrestin1
Components
  • Complex: 5-HT2B receptor bound to LSD in complex with beta-arrestin1
    • Protein or peptide: Anti-5HT2BR Fab light chain
    • Protein or peptide: Anti-5HT2BR Fab heavy chain
    • Protein or peptide: 5-hydroxytryptamine receptor 2B
    • Protein or peptide: Isoform 1B of Beta-arrestin-1
    • Protein or peptide: Fab30 heavy chain
    • Protein or peptide: Fab30 light chain
  • Ligand: 2-acetamido-2-deoxy-beta-D-glucopyranose
  • Ligand: (8alpha)-N,N-diethyl-6-methyl-9,10-didehydroergoline-8-carboxamide

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Supramolecule #1: 5-HT2B receptor bound to LSD in complex with beta-arrestin1

SupramoleculeName: 5-HT2B receptor bound to LSD in complex with beta-arrestin1
type: complex / Chimera: Yes / ID: 1 / Parent: 0 / Macromolecule list: #1-#6

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Macromolecule #1: Anti-5HT2BR Fab light chain

MacromoleculeName: Anti-5HT2BR Fab light chain / type: protein_or_peptide / ID: 1 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Mus musculus (house mouse)
Molecular weightTheoretical: 22.719348 KDa
Recombinant expressionOrganism: Spodoptera frugiperda (fall armyworm)
SequenceString: DIVLIQSPAI MSASPGEKVT ITCSASSSVS YMHWFQQKPG TSPKLWIYST SNLASGVPAR FSGSGSGTSY SLTISRMEAE DAATYYCQQ RSSYPLTFGA GTKLEIKRTV AAPSVFIFPP SDEQLKSGTA SVVCLLNNFY PREAKVQWKV DNALQSGNSQ E SVTEQDSK ...String:
DIVLIQSPAI MSASPGEKVT ITCSASSSVS YMHWFQQKPG TSPKLWIYST SNLASGVPAR FSGSGSGTSY SLTISRMEAE DAATYYCQQ RSSYPLTFGA GTKLEIKRTV AAPSVFIFPP SDEQLKSGTA SVVCLLNNFY PREAKVQWKV DNALQSGNSQ E SVTEQDSK DSTYSLSSTL TLSKADYEKH KVYACEVTHQ GLPVTKSFNR G

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Macromolecule #2: Anti-5HT2BR Fab heavy chain

MacromoleculeName: Anti-5HT2BR Fab heavy chain / type: protein_or_peptide / ID: 2 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Mus musculus (house mouse)
Molecular weightTheoretical: 23.48732 KDa
Recombinant expressionOrganism: Spodoptera frugiperda (fall armyworm)
SequenceString: EVQLQQSGPE LVKPGASVKL SCKASGYTFT SSWMHWVKQR PGQGLEWIGN IYPSNGGTNY NERFKSKATL TVDRSSNTAY MQLSSLTSE DSAVYFCARF GSFITTILTT YYNPVDYWGQ GTTLTVSSAS TKGPSVFPLA PSTAALGCLV KDYFPEPVTV S WNSGALTS ...String:
EVQLQQSGPE LVKPGASVKL SCKASGYTFT SSWMHWVKQR PGQGLEWIGN IYPSNGGTNY NERFKSKATL TVDRSSNTAY MQLSSLTSE DSAVYFCARF GSFITTILTT YYNPVDYWGQ GTTLTVSSAS TKGPSVFPLA PSTAALGCLV KDYFPEPVTV S WNSGALTS GVHTFPAVLQ SSGLYSLSSV VTVPASSLGT QTYICNVNHK PSNTKVDKKV

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Macromolecule #3: 5-hydroxytryptamine receptor 2B

MacromoleculeName: 5-hydroxytryptamine receptor 2B / type: protein_or_peptide / ID: 3 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Homo sapiens (human)
Molecular weightTheoretical: 37.722496 KDa
Recombinant expressionOrganism: Spodoptera frugiperda (fall armyworm)
SequenceString: TESIPEEMKQ IVEEQGNKLH WAALLILMVI IPTIGGNTLV ILAVSLEKKL QYATNYFLMS LAVADLLVGL FVMPIALLTI MFEAMWPLP LVLCPAWLFL DVLFSTASIW HLCAISVDRY IAIKKPIQAN QYNSRATAFI KITVVWLISI GIAIPVPIKG I ETDVDNPN ...String:
TESIPEEMKQ IVEEQGNKLH WAALLILMVI IPTIGGNTLV ILAVSLEKKL QYATNYFLMS LAVADLLVGL FVMPIALLTI MFEAMWPLP LVLCPAWLFL DVLFSTASIW HLCAISVDRY IAIKKPIQAN QYNSRATAFI KITVVWLISI GIAIPVPIKG I ETDVDNPN NITCVLTKER FGDFMLFGSL AAFFTPLAIM IVTYFLTIHA LQKVRLLSGS RQTISNLQRA SKVLGIVFFL FL LMWCPFF ITNITLVLCD SCNQTTLQML LEIFVWIGYV SSGVNPLVYT LFNKTFRDAF GRYITCNYRA TKSVKTPMRL RSS TIQ(SEP)(SEP)(SEP) IILLDTL

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Macromolecule #4: Isoform 1B of Beta-arrestin-1

MacromoleculeName: Isoform 1B of Beta-arrestin-1 / type: protein_or_peptide / ID: 4 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Homo sapiens (human)
Molecular weightTheoretical: 41.345305 KDa
Recombinant expressionOrganism: Spodoptera frugiperda (fall armyworm)
SequenceString: GDKGTRVFKK ASPNGKLTVY LGKRDFVDHI DLVDPVDGVV LVDPEYLKER RVYVTLTCAF RYGREDLDVL GLTFRKDLFV ANVQSFPPA PEDKKPLTRL QERLIKKLGE HAYPFTFEIP PNLPCSVTLQ PGPEDTGKAC GVDYEVKAFC AENLEEKIHK R NSVRLVIE ...String:
GDKGTRVFKK ASPNGKLTVY LGKRDFVDHI DLVDPVDGVV LVDPEYLKER RVYVTLTCAF RYGREDLDVL GLTFRKDLFV ANVQSFPPA PEDKKPLTRL QERLIKKLGE HAYPFTFEIP PNLPCSVTLQ PGPEDTGKAC GVDYEVKAFC AENLEEKIHK R NSVRLVIE KVQYAPERPG PQPTAETTRQ FLMSDKPLHL EASLDKEIYY HGEPISVNVH VTNNTNKTVK KIKISVRQYA DI CLFNTAQ YKCPVAMEEA DDTVAPSSTF CKVYTLTPFL ANNREKRGLA LDGKLKHEDT NLASSTLLRE GANREILGII VSY KVKVKL VVSRGGDVAV ELPFTLMHPK PKEEPPHREV PENETPVDTN L

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Macromolecule #5: Fab30 heavy chain

MacromoleculeName: Fab30 heavy chain / type: protein_or_peptide / ID: 5 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Mus musculus (house mouse)
Molecular weightTheoretical: 13.063479 KDa
Recombinant expressionOrganism: Spodoptera frugiperda (fall armyworm)
SequenceString:
VQLVESGGGL VQPGGSLRLS CAASGFNVYS SSIHWVRQAP GKGLRWVASI SSYYGYTYYA DSVKGRFTIS ADTSKNTAYL QMNSLRAED TAVYYCARSR QFWYSGLDYW GQGTLVTVSS

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Macromolecule #6: Fab30 light chain

MacromoleculeName: Fab30 light chain / type: protein_or_peptide / ID: 6 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Mus musculus (house mouse)
Molecular weightTheoretical: 11.50082 KDa
Recombinant expressionOrganism: Spodoptera frugiperda (fall armyworm)
SequenceString:
DIQMTQSPSS LSASVGDRVT ITCRASQSVS SAVAWYQQKP GKAPKLLIYS ASSLYSGVPS RFSGSRSGTD FTLTISSLQP EDFATYYCQ QYKYVPVTFG QGTKVEI

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Macromolecule #7: 2-acetamido-2-deoxy-beta-D-glucopyranose

MacromoleculeName: 2-acetamido-2-deoxy-beta-D-glucopyranose / type: ligand / ID: 7 / Number of copies: 1 / Formula: NAG
Molecular weightTheoretical: 221.208 Da
Chemical component information

ChemComp-NAG:
2-acetamido-2-deoxy-beta-D-glucopyranose / N-Acetylglucosamine

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Macromolecule #8: (8alpha)-N,N-diethyl-6-methyl-9,10-didehydroergoline-8-carboxamide

MacromoleculeName: (8alpha)-N,N-diethyl-6-methyl-9,10-didehydroergoline-8-carboxamide
type: ligand / ID: 8 / Number of copies: 1 / Formula: 7LD
Molecular weightTheoretical: 323.432 Da
Chemical component information

ChemComp-7LD:
(8alpha)-N,N-diethyl-6-methyl-9,10-didehydroergoline-8-carboxamide / Lysergic acid diethylamide

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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

Concentration3.5 mg/mL
BufferpH: 7.5
VitrificationCryogen name: ETHANE / 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 opticsC2 aperture diameter: 70.0 µm / Calibrated magnification: 57050 / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELDBright-field microscopy / Cs: 2.7 mm / Nominal defocus max: 1.8 µm / Nominal defocus min: 0.8 µm
Sample stageSpecimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER
Image recordingFilm or detector model: GATAN K3 BIOQUANTUM (6k x 4k) / Average electron dose: 1.36 e/Å2
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Particle selectionNumber selected: 4415425
CTF correctionSoftware - Name: cryoSPARC (ver. 3.2)
Initial angle assignmentType: ANGULAR RECONSTITUTION / Software - Name: cryoSPARC (ver. 3.2)
Final angle assignmentType: ANGULAR RECONSTITUTION / Software - Name: cryoSPARC (ver. 3.2)
Final reconstructionResolution.type: BY AUTHOR / Resolution: 3.3 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: cryoSPARC (ver. 3.2) / Number images used: 126485

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