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- EMDB-75847: Influenza A virus Hemagglutinin (A/Darwin/6/2021 H3N2) (C3 symmet... -

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Basic information

Entry
Database: EMDB / ID: EMD-75847
TitleInfluenza A virus Hemagglutinin (A/Darwin/6/2021 H3N2) (C3 symmetry) determined using the SPT Labtech chameleon in the presence of 1x SurfACT
Map dataInfluenza A virus Hemagglutinin (A/Darwin/6/2021 H3N2) (C3 symmetry) determined using the SPT Labtech chameleon in the presence of 1x SurfACT
Sample
  • Complex: Influenza A virus Hemagglutinin (A/Darwin/6/2021 H3N2) (C3 symmetry) determined using the SPT Labtech chameleon in the presence of 1x SurfACT
    • Protein or peptide: Hemagglutinin
  • Ligand: water
KeywordsHemagglutinin / Influenza A virus / H3N2 / Viral Entry / Trimer / Glycoprotein / Membrane Fusion / VIRAL PROTEIN
Function / homology
Function and homology information


viral budding from plasma membrane / clathrin-dependent endocytosis of virus by host cell / host cell surface receptor binding / fusion of virus membrane with host plasma membrane / fusion of virus membrane with host endosome membrane / viral envelope / virion attachment to host cell / host cell plasma membrane / virion membrane / membrane
Similarity search - Function
Haemagglutinin, influenzavirus A / Haemagglutinin, HA1 chain, alpha/beta domain superfamily / Haemagglutinin / Haemagglutinin, influenzavirus A/B / Viral capsid/haemagglutinin protein
Similarity search - Domain/homology
Biological speciesInfluenza A virus
Methodsingle particle reconstruction / cryo EM / Resolution: 2.47 Å
AuthorsEnos SE / Cook BD / Rahmani H / Narehood SM / Li Y / Kuschnerus IC / Redford HT / Dukakis P / Ji D / Bachochin MJ ...Enos SE / Cook BD / Rahmani H / Narehood SM / Li Y / Kuschnerus IC / Redford HT / Dukakis P / Ji D / Bachochin MJ / Grotjahn DA / Herzik MA
Funding support United States, 5 items
OrganizationGrant numberCountry
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)5R35GM138206 United States
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)1S10OD032471 United States
Other privateSearle Scholars United States
Other privateCottrell Scholars United States
Other privateGeorge W. and Carol A. Lattimer Faculty Research Fellowship United States
CitationJournal: bioRxiv / Year: 2026
Title: A Surfactant Cocktail Overcomes Air-Water Interface Artifacts in Single-Particle CryoEM.
Authors: Suzanne E Enos / Brian D Cook / Hamidreza Rahmani / Sarah M Narehood / Yizhou Li / Inga C Kuschnerus / Trevor H Redford / Peter Dukakis / Daniel Ji / Maxwell J Bachochin / Danielle A Grotjahn / Mark A Herzik
Abstract: Single-particle cryogenic electron microscopy (cryoEM) is a widely used technique for structure determination of biomacromolecules to near-atomic resolution. Random distributions of these molecules ...Single-particle cryogenic electron microscopy (cryoEM) is a widely used technique for structure determination of biomacromolecules to near-atomic resolution. Random distributions of these molecules in vitrified ice are necessary to accumulate enough two-dimensional views to generate a complete three-dimensional (3-D) reconstruction. However, interactions between the sample and the air-water interface (AWI) that occur during vitrification often bias the views of the sample, a phenomenon termed preferred orientation, limiting our ability to obtain 3-D reconstructions. Surfactants are often used as sample additives to prevent AWI-induced deterioration, but no general strategy exists for surfactant choice, requiring laborious screening for each sample. To circumvent these issues, we developed SurfACT, a cocktail of diverse surfactants with distinct physicochemical properties that limits AWI-dependent sample denaturation and orientation bias, while mitigating individual surfactant-specific drawbacks. Here we demonstrate SurfACT's effectiveness with four proteins plagued by AWI-induced issues, including two species of hemagglutinin (HA), molybdenum-iron protein (MoFeP) from the nitrogenase enzyme, and aldolase. All four samples show drastically improved viewing distribution and map completeness when SurfACT is applied. Cryogenic electron tomography demonstrates that SurfACT redistributes particles from the AWI into the bulk ice, driving signal recovery and inhibiting denaturation. This versatile sample additive minimizes sample-specific screening and expands the capabilities and range of suitable samples for cryoEM.
History
DepositionMar 5, 2026-
Header (metadata) releaseApr 8, 2026-
Map releaseApr 8, 2026-
UpdateApr 8, 2026-
Current statusApr 8, 2026Processing site: RCSB / Status: Released

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Structure visualization

Supplemental images

emd_75847.png

Downloads & links

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Map

FileDownload / File: emd_75847.map.gz / Format: CCP4 / Size: 343 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationInfluenza A virus Hemagglutinin (A/Darwin/6/2021 H3N2) (C3 symmetry) determined using the SPT Labtech chameleon in the presence of 1x SurfACT
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
0.74 Å/pix.
x 448 pix.
= 329.28 Å		0.74 Å/pix.
x 448 pix.
= 329.28 Å		0.74 Å/pix.
x 448 pix.
= 329.28 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 0.735 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.0533
Minimum - Maximum-0.20344427 - 0.3146481
Average (Standard dev.)0.000058860835 (±0.006594917)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions448448448
Spacing448448448
CellA=B=C: 329.28 Å
α=β=γ: 90.0 °

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Supplemental data

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Additional map: Unsharpened map of Influenza A virus Hemagglutinin (A/Darwin/6/2021...

Fileemd_75847_additional_1.map
AnnotationUnsharpened map of Influenza A virus Hemagglutinin (A/Darwin/6/2021 H3N2) (C3 symmetry) determined using the SPT Labtech chameleon in the presence of 1x SurfACT
Projections & Slices
AxesZYX

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Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Additional map: Mask for Influenza A virus Hemagglutinin (A/Darwin/6/2021 H3N2)...

Fileemd_75847_additional_2.map
AnnotationMask for Influenza A virus Hemagglutinin (A/Darwin/6/2021 H3N2) (C3 symmetry) determined using the SPT Labtech chameleon in the presence of 1x SurfACT
Projections & Slices
AxesZYX

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Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Additional map: Phenix RESOLVE modified EM map of Influenza A...

Fileemd_75847_additional_3.map
AnnotationPhenix RESOLVE modified EM map of Influenza A virus Hemagglutinin (A/Darwin/6/2021 H3N2) (C3 symmetry) determined using the SPT Labtech chameleon in the presence of 1x SurfACT
Projections & Slices
AxesZYX

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Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: Half map A of Influenza A virus Hemagglutinin...

Fileemd_75847_half_map_1.map
AnnotationHalf map A of Influenza A virus Hemagglutinin (A/Darwin/6/2021 H3N2) determined using the SPT Labtech chameleon in the presence of 1x SurfACT; C3 symmetry
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: Half map B of Influenza A virus Hemagglutinin...

Fileemd_75847_half_map_2.map
AnnotationHalf map B of Influenza A virus Hemagglutinin (A/Darwin/6/2021 H3N2) determined using the SPT Labtech chameleon in the presence of 1x SurfACT; C3 symmetry
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Sample components

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Entire : Influenza A virus Hemagglutinin (A/Darwin/6/2021 H3N2) (C3 symmet...

EntireName: Influenza A virus Hemagglutinin (A/Darwin/6/2021 H3N2) (C3 symmetry) determined using the SPT Labtech chameleon in the presence of 1x SurfACT
Components
  • Complex: Influenza A virus Hemagglutinin (A/Darwin/6/2021 H3N2) (C3 symmetry) determined using the SPT Labtech chameleon in the presence of 1x SurfACT
    • Protein or peptide: Hemagglutinin
  • Ligand: water

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Supramolecule #1: Influenza A virus Hemagglutinin (A/Darwin/6/2021 H3N2) (C3 symmet...

SupramoleculeName: Influenza A virus Hemagglutinin (A/Darwin/6/2021 H3N2) (C3 symmetry) determined using the SPT Labtech chameleon in the presence of 1x SurfACT
type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1
Source (natural)Organism: Influenza A virus
Molecular weightTheoretical: 185 KDa

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Macromolecule #1: Hemagglutinin

MacromoleculeName: Hemagglutinin / type: protein_or_peptide / ID: 1 / Number of copies: 3 / Enantiomer: LEVO
Source (natural)Organism: Influenza A virus
Molecular weightTheoretical: 63.30502 KDa
Recombinant expressionOrganism: Homo sapiens (human)
SequenceString: MKTIIALSNI LCLVFAQKIP GNDNSTATLC LGHHAVPNGT IVKTITNDRI EVTNATELVQ NSSIGEICGS PHQILDGGNC TLIDALLGD PQCDGFQNKE WDLFVERSRA NSNCYPYDVP DYASLRSLVA SSGTLEFKNE SFNWTGVKQN GTSSACIRGS S SSFFSRLN ...String:
MKTIIALSNI LCLVFAQKIP GNDNSTATLC LGHHAVPNGT IVKTITNDRI EVTNATELVQ NSSIGEICGS PHQILDGGNC TLIDALLGD PQCDGFQNKE WDLFVERSRA NSNCYPYDVP DYASLRSLVA SSGTLEFKNE SFNWTGVKQN GTSSACIRGS S SSFFSRLN WLTSLNNIYP AQNVTMPNKE QFDKLYIWGV HHPDTDKNQI SLFAQSSGRI TVSTKRSQQA VIPNIGSRPR IR DIPSRIS IYWTIVKPGD ILLINSTGNL IAPRGYFKIR SGKSSIMRSD APIGKCKSEC ITPNGSIPND KPFQNVNRIT YGA CPRYVK QSTLKLATGM RNVPEKQTRG IFGAIAGFIE NGWEGMVDGW YGFRHQNSEG RGQAADLKST QAAIDQINGK LNRL IGKTN EKFHQIEKEF SEVEGRVQDL EKYVEDTKID LWSYNAELLV ALENQHTIDL TDSEMNKLFE KTKKQLRENA EDMGN GCFK IYHKCDNACI GSIRNETYDH NVYRDEALNN RFQIKGVGYI PEAPRDGQAY VRKDGEWVLL STFLGSENLY FQGGSH HHH HH

UniProtKB: Hemagglutinin

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Macromolecule #2: water

MacromoleculeName: water / type: ligand / ID: 2 / Number of copies: 1101 / Formula: HOH
Molecular weightTheoretical: 18.015 Da
Chemical component information

ChemComp-HOH:
WATER

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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

Concentration3.7 mg/mL
BufferpH: 7.6
Component:
ConcentrationFormulaName
50.0 mMNH2C(CH2OH)3-HClTris
150.0 mMNaClSodium Chloride
0.05 %w/vC32H58N2O8SCHAPSO
0.0048 %w/vC20H25F13O11FOM
0.25 %w/v(C6.2H10.3O1.35N0.65Na0.35)72Amphipol A8-35
0.06 %w/v(C2H4O)nC12H26OBrij-35
GridModel: Quantifoil Active R1.2/0.8 / Material: COPPER / Support film - Material: CARBON / Support film - topology: HOLEY / Pretreatment - Type: GLOW DISCHARGE / Pretreatment - Time: 20 sec.
VitrificationCryogen name: ETHANE / Chamber humidity: 75 % / Chamber temperature: 298.15 K / Instrument: SPT LABTECH CHAMELEON
Details: Samples were frozen with the SPT Labtech chameleon.

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Electron microscopy

MicroscopeTFS KRIOS
Specialist opticsEnergy filter - Name: TFS Selectris X / Energy filter - Slit width: 10 eV
Image recordingFilm or detector model: TFS FALCON 4i (4k x 4k) / Digitization - Dimensions - Width: 4096 pixel / Digitization - Dimensions - Height: 4096 pixel / Number grids imaged: 1 / Number real images: 1461 / Average exposure time: 5.0 sec. / Average electron dose: 65.0 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Cs: 2.7 mm / Nominal defocus max: 2.5 µm / Nominal defocus min: 1.0 µm / Nominal magnification: 165000
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

CTF correctionSoftware - Name: cryoSPARC (ver. 4.7.1) / Type: PHASE FLIPPING AND AMPLITUDE CORRECTION
Startup modelType of model: OTHER / Details: Ab-Initio Reconstruction
Final reconstructionApplied symmetry - Point group: C3 (3 fold cyclic) / Algorithm: FOURIER SPACE / Resolution.type: BY AUTHOR / Resolution: 2.47 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: cryoSPARC (ver. 4.7.1) / Number images used: 35526
Initial angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: cryoSPARC (ver. 4.7.1)
Final angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: cryoSPARC (ver. 4.7.1)
FSC plot (resolution estimation)

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Atomic model buiding 1

Initial modelChain - Source name: Other / Chain - Initial model type: experimental model / Details: ModelAngelo
RefinementSpace: REAL / Protocol: RIGID BODY FIT / Overall B value: 68.9
Output model

PDB-11mv:
Influenza A virus Hemagglutinin (A/Darwin/6/2021 H3N2) (C3 symmetry) determined using the SPT Labtech chameleon in the presence of 1x SurfACT

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