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- PDB-7ums: Structure of the VP5*/VP8* assembly from the human rotavirus stra... -

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Basic information

Entry
Database: PDB / ID: 7ums
TitleStructure of the VP5*/VP8* assembly from the human rotavirus strain CDC-9 in complex with antibody 41 - Upright conformation
Components
  • (Outer capsid protein ...) x 2
  • Fab 41 heavy chainFragment antigen-binding
  • Fab 41 light chainFragment antigen-binding
  • Intermediate capsid protein VP6
  • Outer capsid glycoprotein VP7
KeywordsVIRUS / Rotavirus / human / CDC-9 / VP4 / VP5* / VP8* / antibody #41 / broadly neutralizing / cryo-EM
Function / homology
Function and homology information


viral intermediate capsid / host cell endoplasmic reticulum lumen / T=13 icosahedral viral capsid / host cell rough endoplasmic reticulum / host cytoskeleton / viral outer capsid / permeabilization of host organelle membrane involved in viral entry into host cell / symbiont entry into host cell via permeabilization of inner membrane / host cell endoplasmic reticulum-Golgi intermediate compartment / membrane => GO:0016020 ...viral intermediate capsid / host cell endoplasmic reticulum lumen / T=13 icosahedral viral capsid / host cell rough endoplasmic reticulum / host cytoskeleton / viral outer capsid / permeabilization of host organelle membrane involved in viral entry into host cell / symbiont entry into host cell via permeabilization of inner membrane / host cell endoplasmic reticulum-Golgi intermediate compartment / membrane => GO:0016020 / host cell surface receptor binding / fusion of virus membrane with host plasma membrane / viral envelope / virion attachment to host cell / host cell plasma membrane / structural molecule activity / membrane / metal ion binding
Similarity search - Function
Rotavirus A/C, major capsid protein VP6 / Rotavirus major capsid protein VP6 / Glycoprotein VP7 / Glycoprotein VP7, domain 1 / Glycoprotein VP7, domain 2 / Glycoprotein VP7 / Virus capsid protein, alpha-helical / Rotavirus VP4 helical domain / Rotavirus VP4 helical domain / Outer capsid protein VP4 ...Rotavirus A/C, major capsid protein VP6 / Rotavirus major capsid protein VP6 / Glycoprotein VP7 / Glycoprotein VP7, domain 1 / Glycoprotein VP7, domain 2 / Glycoprotein VP7 / Virus capsid protein, alpha-helical / Rotavirus VP4 helical domain / Rotavirus VP4 helical domain / Outer capsid protein VP4 / Rotavirus VP4, membrane interaction domain superfamily / Rotavirus VP4, membrane interaction domain / Rotavirus VP4 membrane interaction domain / Viral capsid/haemagglutinin protein / Concanavalin A-like lectin/glucanase domain superfamily
Similarity search - Domain/homology
Intermediate capsid protein VP6 / Outer capsid glycoprotein VP7 / Outer capsid protein VP4
Similarity search - Component
Biological speciesRotavirus
Homo sapiens (human)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 3.5 Å
AuthorsJenni, S. / Zongli, L. / Wang, Y. / Bessey, T. / Salgado, E.N. / Schmidt, A.G. / Greenberg, H.B. / Jiang, B. / Harrison, S.C.
Funding support United States, 1items
OrganizationGrant numberCountry
National Institutes of Health/National Library of Medicine (NIH/NLM)CA13202 United States
CitationJournal: J Virol / Year: 2022
Title: Rotavirus VP4 Epitope of a Broadly Neutralizing Human Antibody Defined by Its Structure Bound with an Attenuated-Strain Virion.
Authors: Simon Jenni / Zongli Li / Yuhuan Wang / Theresa Bessey / Eric N Salgado / Aaron G Schmidt / Harry B Greenberg / Baoming Jiang / Stephen C Harrison /
Abstract: Rotavirus live-attenuated vaccines, both mono- and pentavalent, generate broadly heterotypic protection. B-cells isolated from adults encode neutralizing antibodies, some with affinity for VP5*, that ...Rotavirus live-attenuated vaccines, both mono- and pentavalent, generate broadly heterotypic protection. B-cells isolated from adults encode neutralizing antibodies, some with affinity for VP5*, that afford broad protection in mice. We have mapped the epitope of one such antibody by determining the high-resolution cryo-EM structure of its antigen-binding fragment (Fab) bound to the virion of a candidate vaccine strain, CDC-9. The Fab contacts both the distal end of a VP5* β-barrel domain and the two VP8* lectin-like domains at the tip of a projecting spike. Its interactions with VP8* do not impinge on the likely receptor-binding site, suggesting that the mechanism of neutralization is at a step subsequent to initial attachment. We also examined structures of CDC-9 virions from two different stages of serial passaging. Nearly all the VP4 (cleaved to VP8*/VP5*) spikes on particles from the earlier passage (wild-type isolate) had transitioned from the "upright" conformation present on fully infectious virions to the "reversed" conformation that is probably the end state of membrane insertion, unable to mediate penetration, consistent with the very low infectivity of the wild-type isolate. About half the VP4 spikes were upright on particles from the later passage, which had recovered substantial infectivity but had acquired an attenuated phenotype in neonatal rats. A mutation in VP4 that occurred during passaging appears to stabilize the interface at the apex of the spike and could account for the greater stability of the upright spikes on the late-passage, attenuated isolate. Rotavirus live-attenuated vaccines generate broadly heterotypic protection, and B-cells isolated from adults encode antibodies that are broadly protective in mice. Determining the structural and mechanistic basis of broad protection can contribute to understanding the current limitations of vaccine efficacy in developing countries. The structure of an attenuated human rotavirus isolate (CDC-9) bound with the Fab fragment of a broadly heterotypic protective antibody shows that protection is probably due to inhibition of the conformational transition in the viral spike protein (VP4) critical for viral penetration, rather than to inhibition of receptor binding. A comparison of structures of CDC-9 virus particles at two stages of serial passaging supports a proposed mechanism for initial steps in rotavirus membrane penetration.
History
DepositionApr 7, 2022Deposition site: RCSB / Processing site: RCSB
Revision 1.0Jul 27, 2022Provider: repository / Type: Initial release
Revision 1.1Aug 3, 2022Group: Author supporting evidence / Source and taxonomy / Structure summary
Category: em_entity_assembly / em_entity_assembly_naturalsource ...em_entity_assembly / em_entity_assembly_naturalsource / em_entity_assembly_recombinant / em_virus_entity / entity_src_gen
Item: _em_virus_entity.entity_assembly_id / _entity_src_gen.pdbx_host_org_ncbi_taxonomy_id / _entity_src_gen.pdbx_host_org_scientific_name
Revision 1.2Aug 17, 2022Group: Database references / Category: citation / citation_author
Item: _citation.country / _citation.journal_abbrev ..._citation.country / _citation.journal_abbrev / _citation.journal_id_ASTM / _citation.journal_id_CSD / _citation.journal_id_ISSN / _citation.page_first / _citation.page_last / _citation.pdbx_database_id_DOI / _citation.pdbx_database_id_PubMed / _citation.title / _citation.year / _citation_author.identifier_ORCID / _citation_author.name
Revision 1.3Aug 31, 2022Group: Database references / Category: citation / Item: _citation.journal_volume

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

Downloads & links

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Assembly

Deposited unit
T: Outer capsid protein VP8*
1: Outer capsid protein VP5*
U: Outer capsid protein VP8*
2: Outer capsid protein VP5*
V: Outer capsid protein VP8*
3: Outer capsid protein VP5*
4: Fab 41 heavy chain
5: Fab 41 light chain
6: Fab 41 heavy chain
7: Fab 41 light chain
A: Intermediate capsid protein VP6
B: Intermediate capsid protein VP6
C: Intermediate capsid protein VP6
D: Intermediate capsid protein VP6
E: Intermediate capsid protein VP6
F: Intermediate capsid protein VP6
G: Intermediate capsid protein VP6
H: Intermediate capsid protein VP6
I: Intermediate capsid protein VP6
J: Intermediate capsid protein VP6
K: Intermediate capsid protein VP6
L: Intermediate capsid protein VP6
M: Intermediate capsid protein VP6
N: Intermediate capsid protein VP6
O: Intermediate capsid protein VP6
P: Intermediate capsid protein VP6
Q: Intermediate capsid protein VP6
R: Intermediate capsid protein VP6
a: Outer capsid glycoprotein VP7
b: Outer capsid glycoprotein VP7
c: Outer capsid glycoprotein VP7
d: Outer capsid glycoprotein VP7
e: Outer capsid glycoprotein VP7
f: Outer capsid glycoprotein VP7
g: Outer capsid glycoprotein VP7
h: Outer capsid glycoprotein VP7
i: Outer capsid glycoprotein VP7
j: Outer capsid glycoprotein VP7
k: Outer capsid glycoprotein VP7
l: Outer capsid glycoprotein VP7
m: Outer capsid glycoprotein VP7
n: Outer capsid glycoprotein VP7
o: Outer capsid glycoprotein VP7
p: Outer capsid glycoprotein VP7
q: Outer capsid glycoprotein VP7
r: Outer capsid glycoprotein VP7
hetero molecules


Theoretical massNumber of molelcules
Total (without water)1,849,951154
Polymers1,838,73646
Non-polymers11,214108
Water0
1


  • Idetical with deposited unit
  • defined by author
  • Evidence: electron microscopy
TypeNameSymmetry operationNumber
identity operation1_5551

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Components

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Outer capsid protein ... , 2 types, 6 molecules TUV123

#1: Protein Outer capsid protein VP8*


Mass: 26248.848 Da / Num. of mol.: 3
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Rotavirus / Strain: CDC-9
Gene: VP4, W907_45309gpVP4, W907_45314gpVP4, W907_45315gpVP4, W907_45321gpVP4, W907_45328gpVP4, W907_45346gpVP4
Production host: Chlorocebus aethiops (grivet) / References: UniProt: X4YMN0
#2: Protein Outer capsid protein VP5*


Mass: 59605.254 Da / Num. of mol.: 3
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Rotavirus / Strain: CDC-9
Gene: VP4, W907_45309gpVP4, W907_45314gpVP4, W907_45315gpVP4, W907_45321gpVP4, W907_45328gpVP4, W907_45346gpVP4
Production host: Chlorocebus aethiops (grivet) / References: UniProt: X4YMN0

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Protein , 2 types, 36 molecules ABCDEFGHIJKLMNOPQRabcdefghijkl...

#5: Protein
Intermediate capsid protein VP6


Mass: 44994.965 Da / Num. of mol.: 18
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Rotavirus / Strain: CDC-9 / Gene: VP6 / Production host: Chlorocebus aethiops (grivet) / References: UniProt: A0A223GHC7
#6: Protein
Outer capsid glycoprotein VP7


Mass: 37435.934 Da / Num. of mol.: 18
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Rotavirus / Strain: CDC-9
Gene: VP7, NC78_51708gpVP7, NC78_52349gpVP7, NC78_52352gpVP7, NC78_52353gpVP7, NC78_52354gpVP7, NC78_52356gpVP7, NC78_52357gpVP7b, W907_45360gpVP7, W907_45361gpVP7, W907_45368gpVP7
Production host: Chlorocebus aethiops (grivet) / References: UniProt: B1NP55

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Antibody , 2 types, 4 molecules 4657

#3: Antibody Fab 41 heavy chain / Fragment antigen-binding


Mass: 25386.260 Da / Num. of mol.: 2
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Cell line (production host): HEK293 / Production host: Homo sapiens (human)
#4: Antibody Fab 41 light chain / Fragment antigen-binding


Mass: 23322.598 Da / Num. of mol.: 2
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Cell line (production host): HEK293 / Production host: Homo sapiens (human)

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Sugars , 2 types, 36 molecules

#7: Polysaccharide beta-D-mannopyranose-(1-4)-2-acetamido-2-deoxy-beta-D-glucopyranose-(1-4)-2-acetamido-2-deoxy-beta- ...beta-D-mannopyranose-(1-4)-2-acetamido-2-deoxy-beta-D-glucopyranose-(1-4)-2-acetamido-2-deoxy-beta-D-glucopyranose


Type: oligosaccharide / Mass: 586.542 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
DescriptorTypeProgram
DManpb1-4DGlcpNAcb1-4DGlcpNAcb1-Glycam Condensed SequenceGMML 1.0
WURCS=2.0/2,3,2/[a2122h-1b_1-5_2*NCC/3=O][a1122h-1b_1-5]/1-1-2/a4-b1_b4-c1WURCSPDB2Glycan 1.1.0
[]{[(4+1)][b-D-GlcpNAc]{[(4+1)][b-D-GlcpNAc]{[(4+1)][b-D-Manp]{}}}}LINUCSPDB-CARE
#8: Sugar...
ChemComp-NAG / 2-acetamido-2-deoxy-beta-D-glucopyranose / N-acetyl-beta-D-glucosamine / 2-acetamido-2-deoxy-beta-D-glucose / 2-acetamido-2-deoxy-D-glucose / 2-acetamido-2-deoxy-glucose / N-ACETYL-D-GLUCOSAMINE / N-Acetylglucosamine


Type: D-saccharide, beta linking / Mass: 221.208 Da / Num. of mol.: 35
Source method: isolated from a genetically manipulated source
Formula: C8H15NO6
IdentifierTypeProgram
DGlcpNAcbCONDENSED IUPAC CARBOHYDRATE SYMBOLGMML 1.0
N-acetyl-b-D-glucopyranosamineCOMMON NAMEGMML 1.0
b-D-GlcpNAcIUPAC CARBOHYDRATE SYMBOLPDB-CARE 1.0
GlcNAcSNFG CARBOHYDRATE SYMBOLGMML 1.0

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Non-polymers , 1 types, 72 molecules

#9: Chemical...
ChemComp-CA / CALCIUM ION


Mass: 40.078 Da / Num. of mol.: 72 / Source method: obtained synthetically / Formula: Ca

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Details

Has ligand of interestN

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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

Component
IDNameTypeEntity IDParent-IDSource
1Rotavirus with Fab boundCOMPLEX#1-#60MULTIPLE SOURCES
2RotavirusVIRUS#1-#2, #5-#61RECOMBINANT
3Fab 41Fragment antigen-bindingCOMPLEX#3-#41RECOMBINANT
Source (natural)
IDEntity assembly-IDOrganismNcbi tax-ID
22Rotavirus10912
33Homo sapiens (human)9606
Source (recombinant)
IDEntity assembly-IDOrganismNcbi tax-ID
22Chlorocebus aethiops (grivet)9534
33Homo sapiens (human)9606
Details of virusEmpty: NO / Enveloped: NO / Isolate: STRAIN / Type: VIRION
Buffer solutionpH: 7.4
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: Tecnai Polara / Image courtesy: FEI Company
MicroscopyModel: FEI POLARA 300
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Illumination mode: FLOOD BEAM
Electron lensMode: BRIGHT FIELDBright-field microscopy / Nominal defocus max: 3500 nm / Nominal defocus min: 1000 nm
Image recordingElectron dose: 60 e/Å2 / Film or detector model: GATAN K2 SUMMIT (4k x 4k)

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Processing

Software
NameVersionClassificationNB
phenix.real_space_refine1.18.2_3874refinement
PHENIX1.18.2_3874refinement
CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
3D reconstructionResolution: 3.5 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 422920 / Symmetry type: POINT
RefinementCross valid method: NONE
Stereochemistry target values: GeoStd + Monomer Library + CDL v1.2
Displacement parametersBiso mean: 83.33 Å2
Refine LS restraints
Refine-IDTypeDev idealNumber
ELECTRON MICROSCOPYf_bond_d0.0056121840
ELECTRON MICROSCOPYf_angle_d0.7679165823
ELECTRON MICROSCOPYf_chiral_restr0.046918920
ELECTRON MICROSCOPYf_plane_restr0.005721424
ELECTRON MICROSCOPYf_dihedral_angle_d6.563916455

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