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- PDB-7epp: Recombinant Alfalfa Mosaic virus coat protein virus-like particle... -

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Basic information

Entry
Database: PDB / ID: 7epp
TitleRecombinant Alfalfa Mosaic virus coat protein virus-like particle (rAMV-CP VLP)
ComponentsCapsid protein
KeywordsVIRUS LIKE PARTICLE / alfalfa mosaic virus / virus-like particle
Function / homologyCoat protein, Ilarvirus / Ilarvirus coat protein / T=3 icosahedral viral capsid / viral nucleocapsid / ribonucleoprotein complex / RNA binding / Capsid protein
Function and homology information
Biological speciesAlfalfa mosaic virus
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 2.4 Å
AuthorsJeong, H. / Lee, S.
CitationJournal: Biochem Biophys Res Commun / Year: 2021
Title: Characterization of alfalfa mosaic virus capsid protein using Cryo-EM.
Authors: Hyeongseop Jeong / Youngmin Park / Sooji Song / Kyungmin Min / Jae-Sung Woo / Young-Ho Lee / Eun-Ju Sohn / Sangmin Lee /
Abstract: VLPs are virus-like particles that comprise viral capsid proteins that can self-assemble and mimic the shape and size of real viral particles; however, because they do not contain genetic material ...VLPs are virus-like particles that comprise viral capsid proteins that can self-assemble and mimic the shape and size of real viral particles; however, because they do not contain genetic material they cannot infect host cells. VLPs have great potential as safe drug/vehicle candidates; therefore, they are gaining popularity in the field of preventive medicine and therapeutics. Indeed, extensive studies are underway to examine their role as carriers for immunization and as vehicles for delivery of therapeutic agents. Here, we examined the possibility of developing VLP-utilizing technology based on an efficient VLP production process and high-resolution structural analysis. Nicotiana benthamiana was used as an expression platform to produce the coat protein of the alfalfa mosaic virus (AMV-CP). About 250 mg/kg of rAMV-CP was produced from Nicotiana benthamiana leaves. Structural analysis revealed that the oligomeric status of rAMV-CP changed according to the composition and pH of the buffer. Size exclusion chromatography and electron microscopy analysis confirmed the optimal conditions for rAMV-CP VLP formation, and a 2.4 Å resolution structure was confirmed by cryo-EM analysis. Based on the efficient protein production, VLP manufacturing technology, and high-resolution structure presented herein, we suggest that rAMV-CP VLP is a useful platform for development of various new drugs.
History
DepositionApr 27, 2021Deposition site: PDBJ / Processing site: PDBJ
Revision 1.0Nov 17, 2021Provider: repository / Type: Initial release
Revision 1.1Jun 5, 2024Group: Data collection / Derived calculations
Category: chem_comp_atom / chem_comp_bond / pdbx_struct_oper_list
Item: _pdbx_struct_oper_list.name / _pdbx_struct_oper_list.symmetry_operation / _pdbx_struct_oper_list.type

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Structure visualization

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  • Biological unit as complete icosahedral assembly
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  • Biological unit as icosahedral pentamer
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  • Biological unit as icosahedral 23 hexamer
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  • Deposited structure unit
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  • Simplified surface model + fitted atomic model
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Assembly

Deposited unit
a: Capsid protein


Theoretical massNumber of molelcules
Total (without water)28,3731
Polymers28,3731
Non-polymers00
Water54030
1
a: Capsid protein
x 60


Theoretical massNumber of molelcules
Total (without water)1,702,40060
Polymers1,702,40060
Non-polymers00
Water1,08160
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1
point symmetry operation59
2


  • Idetical with deposited unit
  • icosahedral asymmetric unit
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1
3
a: Capsid protein
x 5


  • icosahedral pentamer
  • 142 kDa, 5 polymers
Theoretical massNumber of molelcules
Total (without water)141,8675
Polymers141,8675
Non-polymers00
Water905
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1
point symmetry operation4
4
a: Capsid protein
x 6


  • icosahedral 23 hexamer
  • 170 kDa, 6 polymers
Theoretical massNumber of molelcules
Total (without water)170,2406
Polymers170,2406
Non-polymers00
Water1086
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1
point symmetry operation5
5


  • Idetical with deposited unit in distinct coordinate
  • icosahedral asymmetric unit, std point frame
TypeNameSymmetry operationNumber
transform to point frame1
SymmetryPoint symmetry: (Schoenflies symbol: I (icosahedral))

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Components

#1: Protein Capsid protein / CP / Coat protein


Mass: 28373.334 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Alfalfa mosaic virus (strain 425 / isolate Madison)
Gene: ORF3b / Production host: Nicotiana benthamiana (plant) / References: UniProt: P05673
#2: Water ChemComp-HOH / water


Mass: 18.015 Da / Num. of mol.: 30 / Source method: isolated from a natural source / Formula: H2O

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Experimental details

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Experiment

ExperimentMethod: ELECTRON MICROSCOPY
EM experimentAggregation state: PARTICLE / 3D reconstruction method: single particle reconstruction

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Sample preparation

ComponentName: Alfalfa mosaic virus / Type: VIRUS / Entity ID: #1 / Source: RECOMBINANT
Source (natural)Organism: Alfalfa mosaic virus (strain 425 / isolate Madison)
Source (recombinant)Organism: Nicotiana benthamiana (plant)
Details of virusEmpty: YES / Enveloped: NO / Isolate: OTHER / Type: VIRUS-LIKE PARTICLE
Buffer solutionpH: 7
SpecimenEmbedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
VitrificationCryogen name: ETHANE

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Electron microscopy imaging

Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company
MicroscopyModel: FEI TITAN KRIOS
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Illumination mode: FLOOD BEAM
Electron lensMode: BRIGHT FIELD
Image recordingElectron dose: 40 e/Å2 / Detector mode: COUNTING / Film or detector model: FEI FALCON III (4k x 4k)

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Processing

CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
3D reconstructionResolution: 2.4 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 279755 / Symmetry type: POINT

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