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- EMDB-23640: Cryo-EM structure of the HCMV pentamer bound by antibodies 1-103,... -

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Basic information

Entry
Database: EMDB / ID: EMD-23640
TitleCryo-EM structure of the HCMV pentamer bound by antibodies 1-103, 1-32 and 2-25
Map dataFocused refinement calculated in cryoSPARC and sharpened using DeepEMhancer.
Sample
  • Complex: Quaternary complex of the HCMV pentamer bound by Fabs 1-103, 1-32, and 2-25
    • Complex: HCMV pentamer
      • Protein or peptide: x 4 types
    • Complex: 2-25 Fab
      • Protein or peptide: x 2 types
    • Complex: 1-32 Fab
      • Protein or peptide: x 2 types
    • Complex: 1-103 Fab
      • Protein or peptide: x 2 types
  • Ligand: x 1 types
Function / homology
Function and homology information


host cell Golgi apparatus / entry receptor-mediated virion attachment to host cell / symbiont entry into host cell / fusion of virus membrane with host plasma membrane / viral envelope / host cell plasma membrane / virion membrane / membrane
Similarity search - Function
Herpesvirus UL130, cytomegalovirus / HCMV glycoprotein pUL130 / Cytomegalovirus glycoprotein L / Cytomegalovirus glycoprotein L
Similarity search - Domain/homology
Envelope glycoprotein UL130 / UL128 / Envelope glycoprotein L / UL131A
Similarity search - Component
Biological speciesHuman betaherpesvirus 5 / Homo sapiens (human) / Human cytomegalovirus
Methodsingle particle reconstruction / cryo EM / Resolution: 3.81 Å
AuthorsWrapp D / McLellan JS
CitationJournal: Sci Adv / Year: 2022
Title: Structural basis for HCMV Pentamer recognition by neuropilin 2 and neutralizing antibodies.
Authors: Daniel Wrapp / Xiaohua Ye / Zhiqiang Ku / Hang Su / Harrison G Jones / Nianshuang Wang / Akaash K Mishra / Daniel C Freed / Fengsheng Li / Aimin Tang / Leike Li / Dabbu Kumar Jaijyan / Hua ...Authors: Daniel Wrapp / Xiaohua Ye / Zhiqiang Ku / Hang Su / Harrison G Jones / Nianshuang Wang / Akaash K Mishra / Daniel C Freed / Fengsheng Li / Aimin Tang / Leike Li / Dabbu Kumar Jaijyan / Hua Zhu / Dai Wang / Tong-Ming Fu / Ningyan Zhang / Zhiqiang An / Jason S McLellan /
Abstract: Human cytomegalovirus (HCMV) encodes multiple surface glycoprotein complexes to infect a variety of cell types. The HCMV Pentamer, composed of gH, gL, UL128, UL130, and UL131A, enhances entry into ...Human cytomegalovirus (HCMV) encodes multiple surface glycoprotein complexes to infect a variety of cell types. The HCMV Pentamer, composed of gH, gL, UL128, UL130, and UL131A, enhances entry into epithelial, endothelial, and myeloid cells by interacting with the cell surface receptor neuropilin 2 (NRP2). Despite the critical nature of this interaction, the molecular determinants that govern NRP2 recognition remain unclear. Here, we describe the cryo-EM structure of NRP2 bound to Pentamer. The high-affinity interaction between these proteins is calcium dependent and differs from the canonical carboxyl-terminal arginine (CendR) binding that NRP2 typically uses. We also determine the structures of four neutralizing human antibodies bound to the HCMV Pentamer to define susceptible epitopes. Two of these antibodies compete with NRP2 binding, but the two most potent antibodies recognize a previously unidentified epitope that does not overlap the NRP2-binding site. Collectively, these findings provide a structural basis for HCMV tropism and antibody-mediated neutralization.
History
DepositionMar 17, 2021-
Header (metadata) releaseAug 11, 2021-
Map releaseAug 11, 2021-
UpdateMar 23, 2022-
Current statusMar 23, 2022Processing site: RCSB / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 0.476
  • Imaged by UCSF Chimera
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  • Surface view colored by radius
  • Surface level: 0.476
  • Imaged by UCSF Chimera
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  • Surface view with fitted model
  • Atomic models: PDB-7m30
  • Surface level: 0.476
  • Imaged by UCSF Chimera
  • Download
  • Simplified surface model + fitted atomic model
  • Atomic modelsPDB-7m30
  • Imaged by Jmol
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Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

emd_23640.png

Downloads & links

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Map

FileDownload / File: emd_23640.map.gz / Format: CCP4 / Size: 307.5 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationFocused refinement calculated in cryoSPARC and sharpened using DeepEMhancer.
Voxel sizeX=Y=Z: 1.075 Å
Density
Contour LevelBy AUTHOR: 0.476 / Movie #1: 0.476
Minimum - Maximum-0.0007849878 - 3.054172
Average (Standard dev.)0.0007653058 (±0.021013975)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions432432432
Spacing432432432
CellA=B=C: 464.40002 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z1.0751.0751.075
M x/y/z432432432
origin x/y/z0.0000.0000.000
length x/y/z464.400464.400464.400
α/β/γ90.00090.00090.000
start NX/NY/NZ000
NX/NY/NZ400400400
MAP C/R/S123
start NC/NR/NS000
NC/NR/NS432432432
D min/max/mean-0.0013.0540.001

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Supplemental data

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Additional map: Full map that was later subjected to focused...

Fileemd_23640_additional_1.map
AnnotationFull map that was later subjected to focused refinement to calculate the primary map. Calculated in cryoSPARC and sharpened using DeepEMhancer.
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: Half map A.

Fileemd_23640_half_map_1.map
AnnotationHalf map A.
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: Half map B.

Fileemd_23640_half_map_2.map
AnnotationHalf map B.
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Sample components

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Entire : Quaternary complex of the HCMV pentamer bound by Fabs 1-103, 1-32...

EntireName: Quaternary complex of the HCMV pentamer bound by Fabs 1-103, 1-32, and 2-25
Components
  • Complex: Quaternary complex of the HCMV pentamer bound by Fabs 1-103, 1-32, and 2-25
    • Complex: HCMV pentamer
      • Protein or peptide: Envelope glycoprotein L
      • Protein or peptide: Envelope protein UL128
      • Protein or peptide: Envelope glycoprotein UL130
      • Protein or peptide: UL131A
    • Complex: 2-25 Fab
      • Protein or peptide: 2-25 Fab Heavy Chain
      • Protein or peptide: 2-25 Fab Light Chain
    • Complex: 1-32 Fab
      • Protein or peptide: 1-32 Fab Heavy Chain
      • Protein or peptide: 1-32 Fab Light Chain
    • Complex: 1-103 Fab
      • Protein or peptide: 1-103 Fab Heavy Chain
      • Protein or peptide: 1-103 Fab Light Chain
  • Ligand: 2-acetamido-2-deoxy-beta-D-glucopyranose

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Supramolecule #1: Quaternary complex of the HCMV pentamer bound by Fabs 1-103, 1-32...

SupramoleculeName: Quaternary complex of the HCMV pentamer bound by Fabs 1-103, 1-32, and 2-25
type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1-#10
Molecular weightExperimental: 310 KDa

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Supramolecule #2: HCMV pentamer

SupramoleculeName: HCMV pentamer / type: complex / ID: 2 / Parent: 1 / Macromolecule list: #1-#4
Source (natural)Organism: Human betaherpesvirus 5
Recombinant expressionOrganism: Homo sapiens (human)

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Supramolecule #3: 2-25 Fab

SupramoleculeName: 2-25 Fab / type: complex / ID: 3 / Parent: 1 / Macromolecule list: #5-#6
Source (natural)Organism: Homo sapiens (human)
Recombinant expressionOrganism: Homo sapiens (human)

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Supramolecule #4: 1-32 Fab

SupramoleculeName: 1-32 Fab / type: complex / ID: 4 / Parent: 1 / Macromolecule list: #7-#8
Source (natural)Organism: Homo sapiens (human)
Recombinant expressionOrganism: Homo sapiens (human)

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Supramolecule #5: 1-103 Fab

SupramoleculeName: 1-103 Fab / type: complex / ID: 5 / Parent: 1 / Macromolecule list: #9-#10
Source (natural)Organism: Homo sapiens (human)
Recombinant expressionOrganism: Homo sapiens (human)

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Macromolecule #1: Envelope glycoprotein L

MacromoleculeName: Envelope glycoprotein L / type: protein_or_peptide / ID: 1 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Human cytomegalovirus
Molecular weightTheoretical: 27.664645 KDa
Recombinant expressionOrganism: Homo sapiens (human)
SequenceString: VAVSVAPTAA EKVPAECPEL TRRCLLGEVF QGDKYESWLR PLVNVTRRDG PLSQLIRYRP VTPEAANSVL LDDAFLDTLA LLYNNPDQL RALLTLLSSD TAPRWMTVMR GYSECGDGSP AVYTCVDDLC RGYDLTRLSY GRSIFTEHVL GFELVPPSLF N VVVAIRNE ...String:
VAVSVAPTAA EKVPAECPEL TRRCLLGEVF QGDKYESWLR PLVNVTRRDG PLSQLIRYRP VTPEAANSVL LDDAFLDTLA LLYNNPDQL RALLTLLSSD TAPRWMTVMR GYSECGDGSP AVYTCVDDLC RGYDLTRLSY GRSIFTEHVL GFELVPPSLF N VVVAIRNE ATRTNRAVRL PVSTAAAPEG ITLFYGLYNA VKEFCLRHQL DPPLLRHLDK YYAGLPPELK QTRVNLPAHS RY GPQAVDA R

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Macromolecule #2: Envelope protein UL128

MacromoleculeName: Envelope protein UL128 / type: protein_or_peptide / ID: 2 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Human cytomegalovirus
Molecular weightTheoretical: 16.684299 KDa
Recombinant expressionOrganism: Homo sapiens (human)
SequenceString:
EECCEFINVN HPPERCYDFK MCNRFTVALR CPDGEVCYSP EKTAEIRGIV TTMTHSLTRQ VVHNKLTSCN YNPLYLEADG RIRCGKVND KAQYLLGAAG SVPYRWINLE YDKITRIVGL DQYLESVKKH KRLDVCRAKM GYMLQ

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Macromolecule #3: Envelope glycoprotein UL130

MacromoleculeName: Envelope glycoprotein UL130 / type: protein_or_peptide / ID: 3 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Human cytomegalovirus
Molecular weightTheoretical: 21.761678 KDa
Recombinant expressionOrganism: Homo sapiens (human)
SequenceString:
SPWSTLTANQ NPSPPWSKLT YSKPHDAATF YCPFLYPSPP RSPLQFSGFQ QVSTGPECRN ETLYLLYNRE GQTLVERSST WVKKVIWYL SGRNQTILQR MPQTASKPSD GNVQISVEDA KIFGAHMVPK QTKLLRFVVN DGTRYQMCVM KLESWAHVFR D YSVSFQVR LTFTEANNQT YTFCTHPNLI V

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Macromolecule #4: UL131A

MacromoleculeName: UL131A / type: protein_or_peptide / ID: 4 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Human cytomegalovirus
Molecular weightTheoretical: 13.005457 KDa
Recombinant expressionOrganism: Homo sapiens (human)
SequenceString:
QCQRETAEKN DYYRVPHYWD ACSRALPDQT RYKYVEQLVD LTLNYHYDAS HGLDNFDVLK RINVTEVSLL ISDFRRQNRR GGTNKRTTF NAAGSLAPHA RSLEFSVRLF AN

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Macromolecule #5: 2-25 Fab Heavy Chain

MacromoleculeName: 2-25 Fab Heavy Chain / type: protein_or_peptide / ID: 5 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Homo sapiens (human)
Molecular weightTheoretical: 25.206262 KDa
Recombinant expressionOrganism: Homo sapiens (human)
SequenceString: QVQLVESGAE VKKPGASVKV SCKASGYTFT NYAIHWVRQA PGQRLEWMGW INAGRGNTKY SQKFQGRVTI TRDTSASTAY MELSSLRSE DAAVYFCARD ESTGDYYYYM DVWGKGTTVT VSSASTKGPS VFPLAPSSKS TSGGTAALGC LVKDYFPEPV T VSWNSGAL ...String:
QVQLVESGAE VKKPGASVKV SCKASGYTFT NYAIHWVRQA PGQRLEWMGW INAGRGNTKY SQKFQGRVTI TRDTSASTAY MELSSLRSE DAAVYFCARD ESTGDYYYYM DVWGKGTTVT VSSASTKGPS VFPLAPSSKS TSGGTAALGC LVKDYFPEPV T VSWNSGAL TSGVHTFPAV LQSSGLYSLS SVVTVPSSSL GTQTYICNVN HKPSNTKVDK KVEPKSCDKG LEVLFQ

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Macromolecule #6: 2-25 Fab Light Chain

MacromoleculeName: 2-25 Fab Light Chain / type: protein_or_peptide / ID: 6 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Homo sapiens (human)
Molecular weightTheoretical: 22.723951 KDa
Recombinant expressionOrganism: Homo sapiens (human)
SequenceString: SYELTQPPSV SVSPGQTASI TCSGDRLDDK YASWYQQKAG QSPVLVIYQD NKRPSGIPER FSGSNSGNTA TLTISGTQAM DEADYYCQA WDSDTYVFGT GTKVTVLGQP KAAPSVTLFP PSSEELQANK ATLVCLISDF YPGAVTVAWK ADSSPVKAGV E TTTPSKQS ...String:
SYELTQPPSV SVSPGQTASI TCSGDRLDDK YASWYQQKAG QSPVLVIYQD NKRPSGIPER FSGSNSGNTA TLTISGTQAM DEADYYCQA WDSDTYVFGT GTKVTVLGQP KAAPSVTLFP PSSEELQANK ATLVCLISDF YPGAVTVAWK ADSSPVKAGV E TTTPSKQS NNKYAASSYL SLTPEQWKSH KSYSCQVTHE GSTVEKTVAP TECS

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Macromolecule #7: 1-32 Fab Heavy Chain

MacromoleculeName: 1-32 Fab Heavy Chain / type: protein_or_peptide / ID: 7 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Homo sapiens (human)
Molecular weightTheoretical: 26.082371 KDa
Recombinant expressionOrganism: Homo sapiens (human)
SequenceString: QVKLVESGGG VVQPGRSLRL SCAGSGFAFD NYAMHWVRQA PGKGLEWVAV ISLEGRNKYY AGPAKGRFSI SRDNSRNTVH LQMNSLRPE DTAVYFCARD MRYYYDSNGH YRNRYGMDVW GQGTTVIVSS ASTKGPSVFP LAPSSKSTSG GTAALGCLVK D YFPEPVTV ...String:
QVKLVESGGG VVQPGRSLRL SCAGSGFAFD NYAMHWVRQA PGKGLEWVAV ISLEGRNKYY AGPAKGRFSI SRDNSRNTVH LQMNSLRPE DTAVYFCARD MRYYYDSNGH YRNRYGMDVW GQGTTVIVSS ASTKGPSVFP LAPSSKSTSG GTAALGCLVK D YFPEPVTV SWNSGALTSG VHTFPAVLQS SGLYSLSSVV TVPSSSLGTQ TYICNVNHKP SNTKVDKKVE PKSCDKGLEV LF Q

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Macromolecule #8: 1-32 Fab Light Chain

MacromoleculeName: 1-32 Fab Light Chain / type: protein_or_peptide / ID: 8 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Homo sapiens (human)
Molecular weightTheoretical: 23.319729 KDa
Recombinant expressionOrganism: Homo sapiens (human)
SequenceString: DIQMTQSPSS LSASVGDSVT ITCQASQDIN QFVSWYQQKP GKPPKLLIYD ASNLESGVPS RFSGSGSGTH FTFTISSLQP DDIATYYCQ QYENLFTFGP GTKVDIKRTV AAPSVFIFPP SDEQLKSGTA SVVCLLNNFY PREAKVQWKV DNALQSGNSQ E SVTEQDSK ...String:
DIQMTQSPSS LSASVGDSVT ITCQASQDIN QFVSWYQQKP GKPPKLLIYD ASNLESGVPS RFSGSGSGTH FTFTISSLQP DDIATYYCQ QYENLFTFGP GTKVDIKRTV AAPSVFIFPP SDEQLKSGTA SVVCLLNNFY PREAKVQWKV DNALQSGNSQ E SVTEQDSK DSTYSLSSTL TLSKADYEKH KVYACEVTHQ GLSSPVTKSF NRGEC

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Macromolecule #9: 1-103 Fab Heavy Chain

MacromoleculeName: 1-103 Fab Heavy Chain / type: protein_or_peptide / ID: 9 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Homo sapiens (human)
Molecular weightTheoretical: 24.679629 KDa
Recombinant expressionOrganism: Homo sapiens (human)
SequenceString: QLQLQESGPG LVKPSETLSL TCSVSGDAIS GSNYYWGWIR QPPGKGLQWI GSIYHTGSTF YNPSFSSRVT LSVDTSKNQF SLKLISVNA ADTAVYYCAR RIRGYSGTYD WGQGTLVTVS SASTKGPSVF PLAPSSKSTS GGTAALGCLV KDYFPEPVTV S WNSGALTS ...String:
QLQLQESGPG LVKPSETLSL TCSVSGDAIS GSNYYWGWIR QPPGKGLQWI GSIYHTGSTF YNPSFSSRVT LSVDTSKNQF SLKLISVNA ADTAVYYCAR RIRGYSGTYD WGQGTLVTVS SASTKGPSVF PLAPSSKSTS GGTAALGCLV KDYFPEPVTV S WNSGALTS GVHTFPAVLQ SSGLYSLSSV VTVPSSSLGT QTYICNVNHK PSNTKVDKKV EPKSCDKGLE VLFQ

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Macromolecule #10: 1-103 Fab Light Chain

MacromoleculeName: 1-103 Fab Light Chain / type: protein_or_peptide / ID: 10 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Homo sapiens (human)
Molecular weightTheoretical: 22.839244 KDa
Recombinant expressionOrganism: Homo sapiens (human)
SequenceString: DIQLTQSPSF LSASVGDRVT ITCRASQDIS SYVAWYQQKP GNAPKLLISS ASTLPSGVPS RFSGSRSGTD FTLTISSLQP EDFATYYCQ QLNNFGPGTT VDIKRTVAAP SVFIFPPSDE QLKSGTASVV CLLNNFYPRE AKVQWKVDNA LQSGNSQESV T EQDSKDST ...String:
DIQLTQSPSF LSASVGDRVT ITCRASQDIS SYVAWYQQKP GNAPKLLISS ASTLPSGVPS RFSGSRSGTD FTLTISSLQP EDFATYYCQ QLNNFGPGTT VDIKRTVAAP SVFIFPPSDE QLKSGTASVV CLLNNFYPRE AKVQWKVDNA LQSGNSQESV T EQDSKDST YSLSSTLTLS KADYEKHKVY ACEVTHQGLS SPVTKSFNRG EC

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Macromolecule #11: 2-acetamido-2-deoxy-beta-D-glucopyranose

MacromoleculeName: 2-acetamido-2-deoxy-beta-D-glucopyranose / type: ligand / ID: 11 / Number of copies: 2 / Formula: NAG
Molecular weightTheoretical: 221.208 Da
Chemical component information

ChemComp-NAG:
2-acetamido-2-deoxy-beta-D-glucopyranose

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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

Concentration0.2 mg/mL
BufferpH: 8
Component:
ConcentrationFormulaName
2.0 mMC4H11NO3Tris
200.0 mMNaClsodium chloride
0.02 %NaN3sodium azide
0.01 %(C6.2H10.3O1.35N0.65)35amphipol A8-35
GridModel: C-flat-1.2/1.3 / Material: COPPER / Support film - Material: CARBON / Support film - topology: HOLEY ARRAY / Pretreatment - Type: PLASMA CLEANING
VitrificationCryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 277.15 K / Instrument: FEI VITROBOT MARK IV
Details: Blotted for six seconds with a force of "1" before plunging.

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Image recordingFilm or detector model: GATAN K2 SUMMIT (4k x 4k) / Detector mode: COUNTING / Average electron dose: 36.0 e/Å2
Details: Collected from a single grid at a mixture of 0 degrees tilt and -30 degrees tilt
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Particle selectionNumber selected: 834092
CTF correctionSoftware - Name: Warp
Final reconstructionApplied symmetry - Point group: C1 (asymmetric) / Resolution.type: BY AUTHOR / Resolution: 3.81 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: cryoSPARC / Number images used: 198946
Initial angle assignmentType: RANDOM ASSIGNMENT / Software - Name: cryoSPARC
Final angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: cryoSPARC
FSC plot (resolution estimation)

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