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- EMDB-24506: Structure of US27-Gi-scFv16 in CL-state -

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Basic information

Entry
Database: EMDB / ID: EMD-24506
TitleStructure of US27-Gi-scFv16 in CL-state
Map dataFull local anisotropic sharpen map (Phenix).
Sample
  • Complex: US27-Gi-scFv16 complex
    • Complex: Gi heterotrimer
      • Protein or peptide: Guanine nucleotide-binding protein G(i) subunit alpha-1
      • 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
    • Complex: scFv16
      • Protein or peptide: Antibody fragment scFv16
    • Complex: US27
      • Protein or peptide: G-protein coupled receptor homolog US27
KeywordsViral GPCR / HCMV / cytomegalovirus / G protein complex / MEMBRANE PROTEIN
Function / homology
Function and homology information


C-C chemokine receptor activity / C-C chemokine binding / T cell migration / Adenylate cyclase inhibitory pathway / positive regulation of protein localization to cell cortex / regulation of cAMP-mediated signaling / D2 dopamine receptor binding / G protein-coupled serotonin receptor binding / regulation of mitotic spindle organization / cellular response to forskolin ...C-C chemokine receptor activity / C-C chemokine binding / T cell migration / Adenylate cyclase inhibitory pathway / positive regulation of protein localization to cell cortex / regulation of cAMP-mediated signaling / D2 dopamine receptor binding / G protein-coupled serotonin receptor binding / regulation of mitotic spindle organization / cellular response to forskolin / adenylate cyclase-inhibiting G protein-coupled receptor signaling pathway / cell chemotaxis / Regulation of insulin secretion / G protein-coupled receptor binding / calcium-mediated signaling / virion component / G-protein beta/gamma-subunit complex binding / Olfactory Signaling Pathway / Activation of the phototransduction cascade / adenylate cyclase-modulating G protein-coupled receptor signaling pathway / G beta:gamma signalling through PLC beta / Presynaptic function of Kainate receptors / Thromboxane signalling through TP receptor / G protein-coupled acetylcholine receptor signaling pathway / G-protein activation / Activation of G protein gated Potassium channels / Inhibition of voltage gated Ca2+ channels via Gbeta/gamma subunits / Prostacyclin signalling through prostacyclin receptor / Glucagon signaling in metabolic regulation / response to peptide hormone / G beta:gamma signalling through CDC42 / G beta:gamma signalling through BTK / ADP signalling through P2Y purinoceptor 12 / Sensory perception of sweet, bitter, and umami (glutamate) taste / Synthesis, secretion, and inactivation of Glucagon-like Peptide-1 (GLP-1) / photoreceptor disc membrane / Glucagon-type ligand receptors / Adrenaline,noradrenaline inhibits insulin secretion / Vasopressin regulates renal water homeostasis via Aquaporins / G alpha (z) signalling events / Glucagon-like Peptide-1 (GLP1) regulates insulin secretion / cellular response to catecholamine stimulus / ADORA2B mediated anti-inflammatory cytokines production / sensory perception of taste / ADP signalling through P2Y purinoceptor 1 / G beta:gamma signalling through PI3Kgamma / adenylate cyclase-activating dopamine receptor signaling pathway / Cooperation of PDCL (PhLP1) and TRiC/CCT in G-protein beta folding / GPER1 signaling / GDP binding / cellular response to prostaglandin E stimulus / Inactivation, recovery and regulation of the phototransduction cascade / G-protein beta-subunit binding / heterotrimeric G-protein complex / G alpha (12/13) signalling events / extracellular vesicle / signaling receptor complex adaptor activity / Thrombin signalling through proteinase activated receptors (PARs) / GTPase binding / retina development in camera-type eye / Ca2+ pathway / phospholipase C-activating G protein-coupled receptor signaling pathway / positive regulation of cytosolic calcium ion concentration / cell cortex / midbody / fibroblast proliferation / G alpha (i) signalling events / G alpha (s) signalling events / Hydrolases; Acting on acid anhydrides; Acting on GTP to facilitate cellular and subcellular movement / G alpha (q) signalling events / Ras protein signal transduction / cell population proliferation / Extra-nuclear estrogen signaling / immune response / G protein-coupled receptor signaling pathway / external side of plasma membrane / cell division / lysosomal membrane / GTPase activity / virus-mediated perturbation of host defense response / centrosome / synapse / protein-containing complex binding / nucleolus / GTP binding / host cell plasma membrane / magnesium ion binding / signal transduction / extracellular exosome / nucleoplasm / membrane / plasma membrane / cytosol / cytoplasm
Similarity search - Function
: / G-protein alpha subunit, group I / Guanine nucleotide binding protein (G-protein), alpha subunit / G protein alpha subunit, helical insertion / G-protein alpha subunit / G-alpha domain profile. / G protein alpha subunit / G-protein, gamma subunit / G-protein gamma subunit domain profile. / G-protein gamma-like domain ...: / G-protein alpha subunit, group I / Guanine nucleotide binding protein (G-protein), alpha subunit / G protein alpha subunit, helical insertion / G-protein alpha subunit / G-alpha domain profile. / G protein alpha subunit / G-protein, gamma subunit / G-protein gamma subunit domain profile. / G-protein gamma-like domain / G-protein gamma-like domain superfamily / GGL domain / G protein gamma subunit-like motifs / GGL domain / Guanine nucleotide-binding protein, beta subunit / G-protein, beta subunit / G-protein coupled receptors family 1 signature. / G protein-coupled receptor, rhodopsin-like / GPCR, rhodopsin-like, 7TM / G-protein coupled receptors family 1 profile. / 7 transmembrane receptor (rhodopsin family) / G-protein beta WD-40 repeat / WD40 repeat, conserved site / Trp-Asp (WD) repeats signature. / Trp-Asp (WD) repeats profile. / Trp-Asp (WD) repeats circular profile. / WD domain, G-beta repeat / WD40 repeats / WD40 repeat / WD40-repeat-containing domain superfamily / WD40/YVTN repeat-like-containing domain superfamily / P-loop containing nucleoside triphosphate hydrolase
Similarity search - Domain/homology
G-protein coupled receptor homolog US27 / 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(i) subunit alpha-1
Similarity search - Component
Biological speciesHomo sapiens (human) / Mus musculus (house mouse) / Human betaherpesvirus 5 / Human cytomegalovirus
Methodsingle particle reconstruction / cryo EM / Resolution: 3.1 Å
AuthorsTsutsumi N / Jude KM
Funding support United States, 2 items
OrganizationGrant numberCountry
National Institutes of Health/National Institute Of Allergy and Infectious Diseases (NIH/NIAID)R01AI125320 United States
Howard Hughes Medical Institute (HHMI) United States
CitationJournal: Sci Adv / Year: 2022
Title: Atypical structural snapshots of human cytomegalovirus GPCR interactions with host G proteins
Authors: Tsutsumi N / Maeda S / Qu Q / Voegele M / Jude KM / Suomivuori CM / Panova O / Waghray D / Kato HE / Velasco A / Dror RO / Skiniotis G / Kobilka BK / Garcia KC
History
DepositionJul 22, 2021-
Header (metadata) releaseJan 26, 2022-
Map releaseJan 26, 2022-
UpdateNov 13, 2024-
Current statusNov 13, 2024Processing site: RCSB / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 0.12
  • Imaged by UCSF Chimera
  • Download
  • Surface view colored by height
  • Surface level: 0.12
  • Imaged by UCSF Chimera
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  • Surface view with fitted model
  • Atomic models: PDB-7rkx
  • Surface level: 0.12
  • Imaged by UCSF Chimera
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Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

Downloads & links

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Map

FileDownload / File: emd_24506.map.gz / Format: CCP4 / Size: 103 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationFull local anisotropic sharpen map (Phenix).
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesX (Sec.)Y (Row.)Z (Col.)
0.87 Å/pix.
x 300 pix.
= 260.31 Å
0.87 Å/pix.
x 300 pix.
= 260.31 Å
0.87 Å/pix.
x 300 pix.
= 260.31 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 0.8677 Å
Density
Contour LevelBy AUTHOR: 0.11 / Movie #1: 0.12
Minimum - Maximum-0.75623965 - 1.1602594
Average (Standard dev.)0.0009166119 (±0.02798417)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderZYX
Origin000
Dimensions300300300
Spacing300300300
CellA=B=C: 260.31 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z0.86770.86770.8677
M x/y/z300300300
origin x/y/z0.0000.0000.000
length x/y/z260.310260.310260.310
α/β/γ90.00090.00090.000
start NX/NY/NZ000
NX/NY/NZ300300300
MAP C/R/S321
start NC/NR/NS000
NC/NR/NS300300300
D min/max/mean-0.7561.1600.001

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Supplemental data

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Additional map: Full uniform sharpen map used for refinement.

Fileemd_24506_additional_1.map
AnnotationFull uniform sharpen map used for refinement.
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

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Additional map: Full unsharpen map.

Fileemd_24506_additional_2.map
AnnotationFull unsharpen map.
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

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Half map: Half map B.

Fileemd_24506_half_map_1.map
AnnotationHalf map B.
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

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Half map: Half map A.

Fileemd_24506_half_map_2.map
AnnotationHalf map A.
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

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Sample components

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Entire : US27-Gi-scFv16 complex

EntireName: US27-Gi-scFv16 complex
Components
  • Complex: US27-Gi-scFv16 complex
    • Complex: Gi heterotrimer
      • Protein or peptide: Guanine nucleotide-binding protein G(i) subunit alpha-1
      • 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
    • Complex: scFv16
      • Protein or peptide: Antibody fragment scFv16
    • Complex: US27
      • Protein or peptide: G-protein coupled receptor homolog US27

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Supramolecule #1: US27-Gi-scFv16 complex

SupramoleculeName: US27-Gi-scFv16 complex / type: complex / ID: 1 / Parent: 0 / Macromolecule list: all

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Supramolecule #2: Gi heterotrimer

SupramoleculeName: Gi heterotrimer / type: complex / ID: 2 / Parent: 1 / Macromolecule list: #1-#3
Source (natural)Organism: Homo sapiens (human) / Location in cell: Membrane
Molecular weightTheoretical: 90 KDa

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Supramolecule #3: scFv16

SupramoleculeName: scFv16 / type: complex / ID: 3 / Parent: 1 / Macromolecule list: #4
Source (natural)Organism: Mus musculus (house mouse)
Molecular weightTheoretical: 30 KDa

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Supramolecule #4: US27

SupramoleculeName: US27 / type: complex / ID: 4 / Parent: 1 / Macromolecule list: #5
Source (natural)Organism: Human betaherpesvirus 5
Molecular weightTheoretical: 40 KDa

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Macromolecule #1: Guanine nucleotide-binding protein G(i) subunit alpha-1

MacromoleculeName: Guanine nucleotide-binding protein G(i) subunit alpha-1
type: protein_or_peptide / ID: 1 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Homo sapiens (human)
Molecular weightTheoretical: 40.283836 KDa
Recombinant expressionOrganism: Trichoplusia ni (cabbage looper)
SequenceString: GCTLSAEDKA AVERSKMIDR NLREDGEKAA REVKLLLLGA GESGKSTIVK QMKIIHEAGY SEEECKQYKA VVYSNTIQSI IAIIRAMGR LKIDFGDSAR ADDARQLFVL AGAAEEGFMT AELAGVIKRL WKDSGVQACF NRSREYQLND SAAYYLNDLD R IAQPNYIP ...String:
GCTLSAEDKA AVERSKMIDR NLREDGEKAA REVKLLLLGA GESGKSTIVK QMKIIHEAGY SEEECKQYKA VVYSNTIQSI IAIIRAMGR LKIDFGDSAR ADDARQLFVL AGAAEEGFMT AELAGVIKRL WKDSGVQACF NRSREYQLND SAAYYLNDLD R IAQPNYIP TQQDVLRTRV KTTGIVETHF TFKDLHFKMF DVGGQRSERK KWIHCFEGVT AIIFCVALSD YDLVLAEDEE MN RMHESMK LFDSICNNKW FTDTSIILFL NKKDLFEEKI KKSPLTICYP EYAGSNTYEE AAAYIQCQFE DLNKRKDTKE IYT HFTCAT DTKNVQFVFD AVTDVIIKNN LKDCGLF

UniProtKB: Guanine nucleotide-binding protein G(i) subunit alpha-1

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Macromolecule #2: 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: 2 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Homo sapiens (human)
Molecular weightTheoretical: 37.728152 KDa
Recombinant expressionOrganism: Trichoplusia ni (cabbage looper)
SequenceString: GPGSSGSELD QLRQEAEQLK NQIRDARKAC ADATLSQITN NIDPVGRIQM RTRRTLRGHL AKIYAMHWGT DSRLLVSASQ DGKLIIWDS YTTNKVHAIP LRSSWVMTCA YAPSGNYVAC GGLDNICSIY NLKTREGNVR VSRELAGHTG YLSCCRFLDD N QIVTSSGD ...String:
GPGSSGSELD QLRQEAEQLK NQIRDARKAC ADATLSQITN NIDPVGRIQM RTRRTLRGHL AKIYAMHWGT DSRLLVSASQ DGKLIIWDS YTTNKVHAIP LRSSWVMTCA YAPSGNYVAC GGLDNICSIY NLKTREGNVR VSRELAGHTG YLSCCRFLDD N QIVTSSGD TTCALWDIET GQQTTTFTGH TGDVMSLSLA PDTRLFVSGA CDASAKLWDV REGMCRQTFT GHESDINAIC FF PNGNAFA TGSDDATCRL FDLRADQELM TYSHDNIICG ITSVSFSKSG RLLLAGYDDF NCNVWDALKA DRAGVLAGHD NRV SCLGVT DDGMAVATGS WDSFLKIWN

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

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Macromolecule #3: 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: 3 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Homo sapiens (human)
Molecular weightTheoretical: 7.432554 KDa
Recombinant expressionOrganism: Trichoplusia ni (cabbage looper)
SequenceString:
ASNNTASIAQ ARKLVEQLKM EANIDRIKVS KAAADLMAYC EAHAKEDPLL TPVPASENPF REKKFFC

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

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Macromolecule #4: Antibody fragment scFv16

MacromoleculeName: Antibody fragment scFv16 / type: protein_or_peptide / ID: 4 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Mus musculus (house mouse)
Molecular weightTheoretical: 27.340482 KDa
Recombinant expressionOrganism: Trichoplusia ni (cabbage looper)
SequenceString: DVQLVESGGG LVQPGGSRKL SCSASGFAFS SFGMHWVRQA PEKGLEWVAY ISSGSGTIYY ADTVKGRFTI SRDDPKNTLF LQMTSLRSE DTAMYYCVRS IYYYGSSPFD FWGQGTTLTV SSGGGGSGGG GSGGGGSDIV MTQATSSVPV TPGESVSISC R SSKSLLHS ...String:
DVQLVESGGG LVQPGGSRKL SCSASGFAFS SFGMHWVRQA PEKGLEWVAY ISSGSGTIYY ADTVKGRFTI SRDDPKNTLF LQMTSLRSE DTAMYYCVRS IYYYGSSPFD FWGQGTTLTV SSGGGGSGGG GSGGGGSDIV MTQATSSVPV TPGESVSISC R SSKSLLHS NGNTYLYWFL QRPGQSPQLL IYRMSNLASG VPDRFSGSGS GTAFTLTISR LEAEDVGVYY CMQHLEYPLT FG AGTKLEL KGSLEVLFQ

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Macromolecule #5: G-protein coupled receptor homolog US27

MacromoleculeName: G-protein coupled receptor homolog US27 / type: protein_or_peptide / ID: 5 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Human cytomegalovirus
Molecular weightTheoretical: 44.451742 KDa
Recombinant expressionOrganism: Homo sapiens (human)
SequenceString: DYKDDDDMTT STNNQTLTQV SNMTNHTLNS TEIYQLFEYT RLGVWLMCIV GTFLNVLVIT TILYYRRKKK SPSDTYICNL AVADLLIVV GLPFFLEYAK HHPKLSREVV CSGLNACFYI CLFAGVCFLI NLSMDRYCVI VWGVELNRVR NNKRATCWVV I FWILAVLM ...String:
DYKDDDDMTT STNNQTLTQV SNMTNHTLNS TEIYQLFEYT RLGVWLMCIV GTFLNVLVIT TILYYRRKKK SPSDTYICNL AVADLLIVV GLPFFLEYAK HHPKLSREVV CSGLNACFYI CLFAGVCFLI NLSMDRYCVI VWGVELNRVR NNKRATCWVV I FWILAVLM GMPHYLMYSH TNNECVGEFA NETSGWFPVF LNTKVNICGY LAPIALMAYT YNRMVRFIIN YVGKWHMQTL HV LLVVVVS FASFWFPFNL ALFLESIRLL AGVYNDTLQN VIIFCLYVGQ FLAYVRACLN PGIYILVGTQ MRKDMWTTLR VFA CCCVKQ EIPYQDIDIE LQKDIQRRAK HTKRTHYDRK NAPMESGEEE FLLSRGAAHH HHHHHH

UniProtKB: G-protein coupled receptor homolog US27

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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 mg/mL
BufferpH: 7.2
Component:
ConcentrationName
10.0 mMHepes-sodium salt
150.0 mMsodium chloride
0.001 % w/vLauryl Maltose Neopentyl Glycol
0.001 % w/vGlyco-diosgenin

Details: additives: 0.01 % w/v fluorinated octyl maltoside or 0.05 % w/v digitonin
GridModel: Quantifoil R1.2/1.3 / Material: GOLD / Mesh: 300 / Pretreatment - Type: GLOW DISCHARGE / Pretreatment - Time: 40 sec. / Pretreatment - Atmosphere: AIR / Pretreatment - Pressure: 0.039 kPa / Details: 15 mA
VitrificationCryogen name: ETHANE / Chamber humidity: 95 % / Chamber temperature: 293 K / Instrument: LEICA EM GP / Details: 3 s blotting before plunging.

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Specialist opticsEnergy filter - Name: GIF Bioquantum / Energy filter - Slit width: 20 eV
Image recordingFilm or detector model: GATAN K3 (6k x 4k) / Detector mode: SUPER-RESOLUTION / Number grids imaged: 3 / Number real images: 12233 / Average electron dose: 73.0 e/Å2
Details: The data from the three grids were also used to reconstruct the OCL-state US27-Gi-scFv16 complex (7RKY).
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsC2 aperture diameter: 100.0 µm / Calibrated magnification: 57624 / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Cs: 2.7 mm / Nominal defocus max: -2.0 µm / Nominal defocus min: -0.8 µm / Nominal magnification: 105000
Sample stageSpecimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Cooling holder cryogen: NITROGEN
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Particle selectionNumber selected: 12254340
Details: Including non-proteinous features. The actual number of intact complex particles was ~1,159,618.
Startup modelType of model: OTHER
Details: Initial model created by cryoSPARC ab-initio reconstruction.
Final reconstructionResolution.type: BY AUTHOR / Resolution: 3.1 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: cryoSPARC (ver. 3.1)
Details: The resolution estimate was inflated due to the better-defined G protein region compared to the receptor region.
Number images used: 268877
Initial angle assignmentType: ANGULAR RECONSTITUTION
Final angle assignmentType: ANGULAR RECONSTITUTION / Software - Name: cryoSPARC (ver. 3.1)
Final 3D classificationSoftware - Name: cryoSPARC (ver. 3.1)

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Atomic model buiding 1

RefinementSpace: REAL / Protocol: FLEXIBLE FIT
Output model

PDB-7rkx:
Structure of US27-Gi-scFv16 in CL-state

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