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- EMDB-55340: Engineering the ADDomer Nanoparticle Vaccine Scaffold for Improve... -

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Entry
Database: EMDB / ID: EMD-55340
TitleEngineering the ADDomer Nanoparticle Vaccine Scaffold for Improved Assembly and Enhanced Stability.
Map dataMap of two interacting CHIMPSELS_S57C ADDomer pentons from masked 3D classification.
Sample
  • Complex: CHIMPSELS S57C ADDomer: An engineered viral like particle derived from penton base protein of Chimpanzee adenovirus Y25.
    • Protein or peptide: Penton protein
KeywordsAdenovirus / Nanoparticle / Disulphide bond / Thermostable. / VIRUS LIKE PARTICLE
Function / homologyAdenovirus penton base protein / Adenovirus penton base protein / T=25 icosahedral viral capsid / endocytosis involved in viral entry into host cell / virion attachment to host cell / host cell nucleus / structural molecule activity / Penton protein
Function and homology information
Biological speciesChimpanzee adenovirus Y25
Methodsingle particle reconstruction / cryo EM / Resolution: 2.61 Å
AuthorsBalchin G / Berger-Schaffitzel C / Berger I
Funding support United Kingdom, European Union, 3 items
OrganizationGrant numberCountry
Biotechnology and Biological Sciences Research Council (BBSRC) United Kingdom
Engineering and Physical Sciences Research Council United Kingdom
European Research Council (ERC)European Union
CitationJournal: ACS Synth Biol / Year: 2026
Title: Engineering the ADDomer Nanoparticle Vaccine Scaffold for Improved Assembly and Enhanced Stability.
Authors: Georgia Balchin / Burak V Kabasakal / Alessandro Strofaldi / Sophie Hall / Charlotte Fletcher / Dora Buzas / Joshua C Bufton / Sathish K N Yadav / Dakang Shen / Frederic Garzoni / H Adrian ...Authors: Georgia Balchin / Burak V Kabasakal / Alessandro Strofaldi / Sophie Hall / Charlotte Fletcher / Dora Buzas / Joshua C Bufton / Sathish K N Yadav / Dakang Shen / Frederic Garzoni / H Adrian Bunzel / Jennifer J McManus / Christiane Schaffitzel / Imre Berger /
Abstract: Virus-like particles (VLPs) are promising platforms for next-generation vaccines due to their ability to present antigens in highly ordered, repetitive geometries emulating pathogen-associated ...Virus-like particles (VLPs) are promising platforms for next-generation vaccines due to their ability to present antigens in highly ordered, repetitive geometries emulating pathogen-associated patterns to elicit potent immune responses. The ADDomer is a synthetic dodecahedral VLP scaffold derived from the penton base protein (PBP) of human adenovirus serotype 3 (Ad3). PBP tolerates insertion of multiple antigenic epitopes in flexible surface-exposed loops, and spontaneously self-assembles into ADDomer nanoparticles, but faces limitations including incomplete assembly and susceptibility to preexisting antihuman adenovirus immunity. Here, we report two complementary engineering strategies to enhance ADDomer robustness. First, we developed a Chimpanzee adenovirus Y25-based ADDomer (CHIMPSELS) to circumvent preexisting antihuman adenovirus immunity, and introduced a point mutation to restore a motif critical for dodecahedron integrity. Second, we introduced targeted intersubunit disulfide bonds to reinforce particle assembly. High-resolution electron cryo-microscopy confirmed the formation of intact dodecahedral particles, revealing that disulfide bonds stabilize distinct conformations of the PBP N-termini. Differential scanning fluorimetry and dynamic light scattering demonstrated thermal stability and elevated aggregation onset temperatures in the disulfide-stabilized ADDomers, providing a scalable assay for screening ADDomer-based VLP constructs for vaccine development. Incorporation of validated immunogenic epitopes, including a SARS-CoV-2 receptor-binding motif segment and the Chikungunya E2EP3 peptide, demonstrated structural integrity and epitope display by the modified scaffolds. Our results establish a versatile, thermostable VLP platform with reduced susceptibility to preexisting immunity, improved particle integrity, and capacity for modular epitope presentation. This work advances the ADDomer toward practical applications in vaccine development and highlights engineering strategies that can be broadly applied to enhance the performance of protein-based VLP vaccines.
History
DepositionOct 10, 2025-
Header (metadata) releaseMar 25, 2026-
Map releaseMar 25, 2026-
UpdateMar 25, 2026-
Current statusMar 25, 2026Processing site: PDBe / Status: Released

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Structure visualization

Supplemental images

emd_55340.png

Downloads & links

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Map

FileDownload / File: emd_55340.map.gz / Format: CCP4 / Size: 229.8 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationMap of two interacting CHIMPSELS_S57C ADDomer pentons from masked 3D classification.
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
1.07 Å/pix.
x 392 pix.
= 419.44 Å		1.07 Å/pix.
x 392 pix.
= 419.44 Å		1.07 Å/pix.
x 392 pix.
= 419.44 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 1.07 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.022
Minimum - Maximum-0.07444384 - 0.14152353
Average (Standard dev.)0.00012863171 (±0.006575431)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions392392392
Spacing392392392
CellA=B=C: 419.44003 Å
α=β=γ: 90.0 °

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Supplemental data

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Mask #1

Fileemd_55340_msk_1.map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: Half map 1 of two penton CHIMPSELS S57C ADDomer map.

Fileemd_55340_half_map_1.map
AnnotationHalf map 1 of two penton CHIMPSELS_S57C ADDomer map.
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: Half map 2 of two penton CHIMPSELS S57C ADDomer map.

Fileemd_55340_half_map_2.map
AnnotationHalf map 2 of two penton CHIMPSELS_S57C ADDomer map.
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Sample components

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Entire : CHIMPSELS S57C ADDomer: An engineered viral like particle derived...

EntireName: CHIMPSELS S57C ADDomer: An engineered viral like particle derived from penton base protein of Chimpanzee adenovirus Y25.
Components
  • Complex: CHIMPSELS S57C ADDomer: An engineered viral like particle derived from penton base protein of Chimpanzee adenovirus Y25.
    • Protein or peptide: Penton protein

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Supramolecule #1: CHIMPSELS S57C ADDomer: An engineered viral like particle derived...

SupramoleculeName: CHIMPSELS S57C ADDomer: An engineered viral like particle derived from penton base protein of Chimpanzee adenovirus Y25.
type: complex / ID: 1 / Parent: 0 / Macromolecule list: all
Details: ADDomer produced by recombinant expression of the engineered penton base protein of Chimpanzee adenovirus Y25 using the MultiBac baculovirus-insect cell expression system. 60 penton base ...Details: ADDomer produced by recombinant expression of the engineered penton base protein of Chimpanzee adenovirus Y25 using the MultiBac baculovirus-insect cell expression system. 60 penton base proteins self assemble into the ADDomer particle.
Source (natural)Organism: Chimpanzee adenovirus Y25
Molecular weightTheoretical: 3.65 MDa

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Macromolecule #1: Penton protein

MacromoleculeName: Penton protein / type: protein_or_peptide / ID: 1 / Number of copies: 10 / Enantiomer: LEVO
Source (natural)Organism: Chimpanzee adenovirus Y25
Molecular weightTheoretical: 60.854137 KDa
Recombinant expressionOrganism: Trichoplusia ni (cabbage looper)
SequenceString: MMRRAYPEGP PPSYESVMQQ AMAAAAAMQP PLEAPYVPPR YLAPTEGRNS IRYSELCPLY DTTRLYLVDN KSADIASLNY QNDHSNFLT TVVQNNDFTP TEASTQTINF DERSRWGGQL KTIMHTNMPN VNEFMYSNKF KARVMVSRKT PNGEFVTVTD G PGSQDILE ...String:
MMRRAYPEGP PPSYESVMQQ AMAAAAAMQP PLEAPYVPPR YLAPTEGRNS IRYSELCPLY DTTRLYLVDN KSADIASLNY QNDHSNFLT TVVQNNDFTP TEASTQTINF DERSRWGGQL KTIMHTNMPN VNEFMYSNKF KARVMVSRKT PNGEFVTVTD G PGSQDILE YEWVEFELPE GNFSVTMTID LMNNAIIDNY LAVGRQNGVL ESDIGVKFDT RNFRLGWDPV TELVMPGVYT NE AFHPDIV LLPGCGVDFT ESRLSNLLGI RKRQPFQEGF QIMYEDLEGG NIPALLDVDA YEKSKEESAA AARTAAVATA STE VDVRGD NFASPAAELV AAAEAAETES SRKIVIQPVE KDSKDRSYNV LPDKINTAYR SWYLAYNYGD PEKGVRSWTL LTTS DVTCG VEQVYWSLPD MMQDPVTFRS TRQVSNYPVV GAELLPVYSK SFFNEQAVYS QQLRAFTSLT HVFNRFPENQ ILVRP PAPT ITTVSENVPA LTDHGTLPLR SSIRGVQRVT VTDARRRTCP YVYKALGIVA PRVLSSRTF

UniProtKB: Penton protein

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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

Concentration0.5 mg/mL
BufferpH: 7.5
Component:
ConcentrationFormulaName
50.0 mMTrisTris
150.0 mMNaClsodium chloride
0.2 mMEDTAEthylenediaminetetraacetic acid
0.1 mMAEBSF4-(2-Aminoethyl)benzenesulfonyl fluoride
100.0 uMDichloro(1,10-phenanthroline)copper(II)Copper Phenanthroline
GridModel: Quantifoil R1.2/1.3 / Material: COPPER / Pretreatment - Type: GLOW DISCHARGE / Pretreatment - Time: 120 sec. / Pretreatment - Atmosphere: AIR / Details: glow discharged at 4 mA for 120 seconds
VitrificationCryogen name: ETHANE-PROPANE / Chamber humidity: 100 % / Chamber temperature: 277.15 K / Instrument: FEI VITROBOT MARK II

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Electron microscopy

MicroscopeTFS KRIOS
Image recordingFilm or detector model: GATAN K3 (6k x 4k) / Number grids imaged: 1 / Number real images: 16511 / Average exposure time: 3.846 sec. / Average electron dose: 50.0 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Nominal defocus max: 2.0 µm / Nominal defocus min: 0.8 µm
Sample stageCooling holder cryogen: NITROGEN
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Particle selectionNumber selected: 2464479
CTF correctionSoftware - Name: Gctf / Type: PHASE FLIPPING AND AMPLITUDE CORRECTION
Startup modelType of model: OTHER
Details: used a 60 angstrom low pass filtered map of the CHIMPSELS ADDomer.
Final reconstructionResolution.type: BY AUTHOR / Resolution: 2.61 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: RELION (ver. 4.0) / Number images used: 113117
Initial angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: RELION (ver. 4.0)
Final angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: RELION (ver. 4.0)
Final 3D classificationSoftware - Name: RELION

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Atomic model buiding 1

Initial modelChain - Source name: Other / Chain - Initial model type: experimental model
Details: The CHIMPSELS ADDomer model was used an initial model source.
RefinementSpace: REAL / Protocol: RIGID BODY FIT
Output model

PDB-9sy5:
Engineering the ADDomer Nanoparticle Vaccine Scaffold for Improved Assembly and Enhanced Stability.

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