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- PDB-8d3a: Crystal Structure of DH475 Fab in complex with Man9 -

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Basic information

Entry
Database: PDB / ID: 8d3a
TitleCrystal Structure of DH475 Fab in complex with Man9
Components
  • DH475 Fab heavy chain
  • DH475 Fab light chain
KeywordsIMMUNE SYSTEM/ANTIGEN / FAB FRAGMENT / HIV-1 / ANTIBODY / IMMUNE SYSTEM / IMMUNE SYSTEM-ANTIGEN complex
Biological speciesHomo sapiens (human)
MethodX-RAY DIFFRACTION / SYNCHROTRON / MOLECULAR REPLACEMENT / Resolution: 2.9 Å
AuthorsFinkelstein, M. / Fera, D.
Funding support United States, 1items
OrganizationGrant numberCountry
National Institutes of Health/National Institute Of Allergy and Infectious Diseases (NIH/NIAID)1R15AI150484 United States
CitationJournal: Front Immunol / Year: 2022
Title: Analysis of two cooperating antibodies unveils immune pressure imposed on HIV Env to elicit a V3-glycan supersite broadly neutralizing antibody lineage.
Authors: Maxwell T Finkelstein / Emma Parker Miller / Molly C Erdman / Daniela Fera /
Abstract: Elicitation of broadly neutralizing antibodies (bnAbs) is a goal of vaccine design as a strategy for targeting highly divergent strains of HIV-1. Current HIV-1 vaccine design efforts seek to elicit ...Elicitation of broadly neutralizing antibodies (bnAbs) is a goal of vaccine design as a strategy for targeting highly divergent strains of HIV-1. Current HIV-1 vaccine design efforts seek to elicit bnAbs by first eliciting their precursors through prime-boost regimens. This requires an understanding of the co-evolution between viruses and antibodies. Towards this goal, we have analyzed two cooperating antibodies, DH475 and DH272, which exerted pressure on the HIV population in an infected donor, called CH848, to evolve in such a way that it became sensitive to the V3-glycan supersite DH270 bnAb lineage. We obtained a 2.90Å crystal structure of DH475 in complex with the Man glycan and a negative stain EM model of DH272 in complex with the HIV-1 spike trimer, Env. Coupled with additional modeling studies and biochemical data, our studies reveal that DH475 contacts a V3- and V4-glycan dependent epitope accessible on an open or shed Env and that DH272 makes critical contacts with the V1V2 and V3 loops on HIV-1 Env. Using these data, we suggest a prime-boost regimen that may facilitate the initiation of DH270-like bnAb precursors.
History
DepositionJun 1, 2022Deposition site: RCSB / Processing site: RCSB
Revision 1.0Oct 12, 2022Provider: repository / Type: Initial release
Revision 1.1Oct 26, 2022Group: Database references / Category: citation
Item: _citation.journal_volume / _citation.page_first ..._citation.journal_volume / _citation.page_first / _citation.page_last / _citation.pdbx_database_id_PubMed / _citation.title
Revision 1.2Oct 18, 2023Group: Data collection / Refinement description
Category: chem_comp_atom / chem_comp_bond / pdbx_initial_refinement_model

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

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Assembly

Deposited unit
H: DH475 Fab heavy chain
L: DH475 Fab light chain
hetero molecules


Theoretical massNumber of molelcules
Total (without water)49,4983
Polymers47,6142
Non-polymers1,8841
Water1,17165
1


  • Idetical with deposited unit
  • defined by author&software
  • Evidence: gel filtration
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1
Buried area6070 Å2
ΔGint18 kcal/mol
Surface area20540 Å2
MethodPISA
Unit cell
Length a, b, c (Å)126.726, 126.726, 215.816
Angle α, β, γ (deg.)90.000, 90.000, 120.000
Int Tables number178
Space group name H-MP6122

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Components

#1: Antibody DH475 Fab heavy chain


Mass: 24447.354 Da / Num. of mol.: 1 / Fragment: FAB
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Cell line: HEK293T / Plasmid: pVRC-8400 / Cell line (production host): HEK293T / Production host: Homo sapiens (human)
#2: Antibody DH475 Fab light chain


Mass: 23166.574 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Cell line (production host): HEK293T / Production host: Homo sapiens (human)
#3: Polysaccharide alpha-D-mannopyranose-(1-2)-alpha-D-mannopyranose-(1-2)-alpha-D-mannopyranose-(1-3)-[alpha-D- ...alpha-D-mannopyranose-(1-2)-alpha-D-mannopyranose-(1-2)-alpha-D-mannopyranose-(1-3)-[alpha-D-mannopyranose-(1-2)-alpha-D-mannopyranose-(1-3)-[alpha-D-mannopyranose-(1-2)-alpha-D-mannopyranose-(1-6)]alpha-D-mannopyranose-(1-6)]beta-D-mannopyranose-(1-4)-2-acetamido-2-deoxy-beta-D-glucopyranose-(1-4)-2-acetamido-2-deoxy-beta-D-glucopyranose


Type: oligosaccharide / Mass: 1883.668 Da / Num. of mol.: 1 / Source method: obtained synthetically
DescriptorTypeProgram
DManpa1-2DManpa1-2DManpa1-3[DManpa1-2DManpa1-3[DManpa1-2DManpa1-6]DManpa1-6]DManpb1-4DGlcpNAcb1-4DGlcpNAcb1-ROHGlycam Condensed SequenceGMML 1.0
WURCS=2.0/3,11,10/[a2122h-1b_1-5_2*NCC/3=O][a1122h-1b_1-5][a1122h-1a_1-5]/1-1-2-3-3-3-3-3-3-3-3/a4-b1_b4-c1_c3-d1_c6-g1_d2-e1_e2-f1_g3-h1_g6-j1_h2-i1_j2-k1WURCSPDB2Glycan 1.1.0
[][D-1-deoxy-GlcpNAc]{[(4+1)][b-D-GlcpNAc]{[(4+1)][b-D-Manp]{[(3+1)][a-D-Manp]{[(2+1)][a-D-Manp]{[(2+1)][a-D-Manp]{}}}[(6+1)][a-D-Manp]{[(3+1)][a-D-Manp]{[(2+1)][a-D-Manp]{}}[(6+1)][a-D-Manp]{[(2+1)][a-D-Manp]{}}}}}}LINUCSPDB-CARE
#4: Water ChemComp-HOH / water


Mass: 18.015 Da / Num. of mol.: 65 / Source method: isolated from a natural source / Formula: H2O
Has ligand of interestY
Nonpolymer detailsThe reducing end NAG was altered to place an amine group on C1, which was then reacted with biotin. ...The reducing end NAG was altered to place an amine group on C1, which was then reacted with biotin. The electron density for the biotin, however, is not clear, and so the biotin is not modeled.

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Experimental details

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Experiment

ExperimentMethod: X-RAY DIFFRACTION / Number of used crystals: 1

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Sample preparation

CrystalDensity Matthews: 5.25 Å3/Da / Density % sol: 76.59 %
Crystal growTemperature: 293 K / Method: vapor diffusion, hanging drop / pH: 6 / Details: 1.5 M (NH4)2SO4, 100 mM PIPES, pH 6.0

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Data collection

DiffractionMean temperature: 100 K / Serial crystal experiment: N
Diffraction sourceSource: SYNCHROTRON / Site: APS / Beamline: 24-ID-C / Wavelength: 0.9792 Å
DetectorType: DECTRIS EIGER2 X 16M / Detector: PIXEL / Date: Aug 3, 2016
RadiationProtocol: SINGLE WAVELENGTH / Monochromatic (M) / Laue (L): M / Scattering type: x-ray
Radiation wavelengthWavelength: 0.9792 Å / Relative weight: 1
ReflectionResolution: 2.9→76.94 Å / Num. obs: 22928 / % possible obs: 98 % / Redundancy: 3.1 % / CC1/2: 0.993 / Net I/σ(I): 8
Reflection shellResolution: 2.9→3.08 Å / Rmerge(I) obs: 1.064 / Num. unique obs: 3672 / % possible all: 99.1

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Processing

Software
NameVersionClassification
XDSdata scaling
PHENIX1.18.2_3874refinement
PDB_EXTRACT3.27data extraction
XDSdata reduction
PHASERphasing
RefinementMethod to determine structure: MOLECULAR REPLACEMENT
Starting model: 5TPP

5tpp


Resolution: 2.9→76.94 Å / SU ML: 0.45 / Cross valid method: THROUGHOUT / σ(F): 1.34 / Phase error: 28.91 / Stereochemistry target values: ML
RfactorNum. reflection% reflection
Rfree0.272 1105 4.84 %
Rwork0.2291 21720 -
obs0.2312 22825 97.43 %
Solvent computationShrinkage radii: 0.9 Å / VDW probe radii: 1.11 Å / Solvent model: FLAT BULK SOLVENT MODEL
Displacement parametersBiso max: 146.16 Å2 / Biso mean: 82.3971 Å2 / Biso min: 46.95 Å2
Refinement stepCycle: final / Resolution: 2.9→76.94 Å
ProteinNucleic acidLigandSolventTotal
Num. atoms3295 0 127 65 3487
Biso mean--81.32 73.24 -
Num. residues----441
LS refinement shell

Refine-ID: X-RAY DIFFRACTION / Rfactor Rfree error: 0 / Total num. of bins used: 8

Resolution (Å)Rfactor RfreeNum. reflection RfreeRfactor RworkNum. reflection RworkNum. reflection all% reflection obs (%)
2.9-3.030.37171240.3522646277097
3.03-3.190.3811450.3382692283799
3.19-3.390.36991320.30612710284299
3.39-3.650.31421500.26082712286299
3.65-4.020.27211330.2352680281397
4.02-4.60.21791390.18582711285097
4.6-5.80.24951260.18542747287397
5.8-76.940.24941560.21822822297894
Refinement TLS params.Method: refined / Origin x: -12.7044 Å / Origin y: -45.7812 Å / Origin z: 14.3291 Å
111213212223313233
T0.6369 Å20.1168 Å2-0.0447 Å2-0.641 Å2-0.1568 Å2--0.4842 Å2
L2.9154 °2-0.6015 °21.1467 °2-0.2573 °2-0.7078 °2--0.0367 °2
S-0.1702 Å °-0.1779 Å °0.0968 Å °-0.1459 Å °0.1326 Å °0.0102 Å °-0.0228 Å °-0.1 Å °0.0319 Å °
Refinement TLS group
IDRefine-IDRefine TLS-IDSelection detailsAuth asym-IDAuth seq-ID
1X-RAY DIFFRACTION1allH1 - 227
2X-RAY DIFFRACTION1allL1 - 214
3X-RAY DIFFRACTION1allG1 - 11
4X-RAY DIFFRACTION1allW1 - 76

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