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- PDB-4jy4: Crystal structure of human Fab PGT121, a broadly reactive and pot... -

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Entry
Database: PDB / ID: 4jy4
TitleCrystal structure of human Fab PGT121, a broadly reactive and potent HIV-1 neutralizing antibody
Components
  • PGT121 heavy chain
  • PGT121 light chain
KeywordsIMMUNE SYSTEM / Broadly neutralizing antibody against HIV-1 / HIV-1 Env gp120 subunit
Function / homologyImmunoglobulins / Immunoglobulin-like / Sandwich / Mainly Beta
Function and homology information
Biological speciesHomo sapiens (human)
MethodX-RAY DIFFRACTION / SYNCHROTRON / MOLECULAR REPLACEMENT / Resolution: 2.8 Å
AuthorsJulien, J.-P. / Diwanji, D.C. / Burton, D.R. / Wilson, I.A.
CitationJournal: PLoS Pathog / Year: 2013
Title: Broadly neutralizing antibody PGT121 allosterically modulates CD4 binding via recognition of the HIV-1 gp120 V3 base and multiple surrounding glycans.
Authors: Jean-Philippe Julien / Devin Sok / Reza Khayat / Jeong Hyun Lee / Katie J Doores / Laura M Walker / Alejandra Ramos / Devan C Diwanji / Robert Pejchal / Albert Cupo / Umesh Katpally / Rafael ...Authors: Jean-Philippe Julien / Devin Sok / Reza Khayat / Jeong Hyun Lee / Katie J Doores / Laura M Walker / Alejandra Ramos / Devan C Diwanji / Robert Pejchal / Albert Cupo / Umesh Katpally / Rafael S Depetris / Robyn L Stanfield / Ryan McBride / Andre J Marozsan / James C Paulson / Rogier W Sanders / John P Moore / Dennis R Burton / Pascal Poignard / Andrew B Ward / Ian A Wilson /
Abstract: New broad and potent neutralizing HIV-1 antibodies have recently been described that are largely dependent on the gp120 N332 glycan for Env recognition. Members of the PGT121 family of antibodies, ...New broad and potent neutralizing HIV-1 antibodies have recently been described that are largely dependent on the gp120 N332 glycan for Env recognition. Members of the PGT121 family of antibodies, isolated from an African donor, neutralize ∼70% of circulating isolates with a median IC50 less than 0.05 µg ml(-1). Here, we show that three family members, PGT121, PGT122 and PGT123, have very similar crystal structures. A long 24-residue HCDR3 divides the antibody binding site into two functional surfaces, consisting of an open face, formed by the heavy chain CDRs, and an elongated face, formed by LCDR1, LCDR3 and the tip of the HCDR3. Alanine scanning mutagenesis of the antibody paratope reveals a crucial role in neutralization for residues on the elongated face, whereas the open face, which accommodates a complex biantennary glycan in the PGT121 structure, appears to play a more secondary role. Negative-stain EM reconstructions of an engineered recombinant Env gp140 trimer (SOSIP.664) reveal that PGT122 interacts with the gp120 outer domain at a more vertical angle with respect to the top surface of the spike than the previously characterized antibody PGT128, which is also dependent on the N332 glycan. We then used ITC and FACS to demonstrate that the PGT121 antibodies inhibit CD4 binding to gp120 despite the epitope being distal from the CD4 binding site. Together, these structural, functional and biophysical results suggest that the PGT121 antibodies may interfere with Env receptor engagement by an allosteric mechanism in which key structural elements, such as the V3 base, the N332 oligomannose glycan and surrounding glycans, including a putative V1/V2 complex biantennary glycan, are conformationally constrained.
History
DepositionMar 29, 2013Deposition site: RCSB / Processing site: RCSB
Revision 1.0May 15, 2013Provider: repository / Type: Initial release
Revision 1.1Feb 5, 2014Group: Database references
Revision 2.0Jul 29, 2020Group: Advisory / Atomic model ...Advisory / Atomic model / Data collection / Derived calculations / Structure summary
Category: atom_site / chem_comp ...atom_site / chem_comp / entity / pdbx_branch_scheme / pdbx_chem_comp_identifier / pdbx_entity_branch / pdbx_entity_branch_descriptor / pdbx_entity_branch_link / pdbx_entity_branch_list / pdbx_entity_nonpoly / pdbx_nonpoly_scheme / pdbx_struct_assembly_gen / pdbx_validate_close_contact / struct_asym / struct_conn / struct_site / struct_site_gen
Item: _atom_site.B_iso_or_equiv / _atom_site.Cartn_x ..._atom_site.B_iso_or_equiv / _atom_site.Cartn_x / _atom_site.Cartn_y / _atom_site.Cartn_z / _atom_site.auth_asym_id / _atom_site.auth_atom_id / _atom_site.auth_comp_id / _atom_site.auth_seq_id / _atom_site.label_asym_id / _atom_site.label_atom_id / _atom_site.label_comp_id / _atom_site.label_entity_id / _atom_site.type_symbol / _chem_comp.name / _chem_comp.type / _pdbx_struct_assembly_gen.asym_id_list / _pdbx_validate_close_contact.auth_asym_id_1 / _pdbx_validate_close_contact.auth_asym_id_2 / _pdbx_validate_close_contact.auth_seq_id_1 / _pdbx_validate_close_contact.auth_seq_id_2 / _struct_conn.pdbx_dist_value / _struct_conn.pdbx_leaving_atom_flag / _struct_conn.pdbx_role / _struct_conn.ptnr1_auth_asym_id / _struct_conn.ptnr1_auth_comp_id / _struct_conn.ptnr1_auth_seq_id / _struct_conn.ptnr1_label_asym_id / _struct_conn.ptnr1_label_atom_id / _struct_conn.ptnr1_label_comp_id / _struct_conn.ptnr1_label_seq_id / _struct_conn.ptnr2_auth_asym_id / _struct_conn.ptnr2_auth_comp_id / _struct_conn.ptnr2_auth_seq_id / _struct_conn.ptnr2_label_asym_id / _struct_conn.ptnr2_label_comp_id
Description: Carbohydrate remediation / Provider: repository / Type: Remediation
Revision 2.1Sep 20, 2023Group: Data collection / Database references ...Data collection / Database references / Refinement description / Structure summary
Category: chem_comp / chem_comp_atom ...chem_comp / chem_comp_atom / chem_comp_bond / database_2 / pdbx_initial_refinement_model
Item: _chem_comp.pdbx_synonyms / _database_2.pdbx_DOI / _database_2.pdbx_database_accession

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

Downloads & links

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Assembly

Deposited unit
B: PGT121 heavy chain
A: PGT121 light chain
hetero molecules


Theoretical massNumber of molelcules
Total (without water)49,7346
Polymers47,7442
Non-polymers1,9904
Water23413
1


  • Idetical with deposited unit
  • defined by author&software
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1
Buried area6320 Å2
ΔGint0 kcal/mol
Surface area20670 Å2
MethodPISA
Unit cell
Length a, b, c (Å)64.970, 65.720, 159.210
Angle α, β, γ (deg.)90.00, 90.00, 90.00
Int Tables number19
Space group name H-MP212121

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Components

#1: Antibody PGT121 heavy chain


Mass: 25169.230 Da / Num. of mol.: 1 / Fragment: Fab
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Cell line (production host): HEK 293F / Production host: Homo sapiens (human)
#2: Antibody PGT121 light chain


Mass: 22574.947 Da / Num. of mol.: 1 / Fragment: Fab
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Cell line (production host): HEK 293F / Production host: Homo sapiens (human)
#3: Polysaccharide N-acetyl-alpha-neuraminic acid-(2-6)-beta-D-galactopyranose-(1-4)-2-acetamido-2-deoxy-beta-D- ...N-acetyl-alpha-neuraminic acid-(2-6)-beta-D-galactopyranose-(1-4)-2-acetamido-2-deoxy-beta-D-glucopyranose-(1-2)-alpha-D-mannopyranose-(1-3)-[alpha-D-mannopyranose-(1-6)]beta-D-mannopyranose-(1-4)-2-acetamido-2-deoxy-beta-D-glucopyranose-(1-4)-[alpha-L-fucopyranose-(1-6)]2-acetamido-2-deoxy-beta-D-glucopyranose


Type: oligosaccharide / Mass: 1713.552 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
DescriptorTypeProgram
DNeup5Aca2-6DGalpb1-4DGlcpNAcb1-2DManpa1-3[DManpa1-6]DManpb1-4DGlcpNAcb1-4[LFucpa1-6]DGlcpNAcb1-Glycam Condensed SequenceGMML 1.0
WURCS=2.0/6,9,8/[a2122h-1b_1-5_2*NCC/3=O][a1122h-1b_1-5][a1122h-1a_1-5][a2112h-1b_1-5][Aad21122h-2a_2-6_5*NCC/3=O][a1221m-1a_1-5]/1-1-2-3-1-4-5-3-6/a4-b1_a6-i1_b4-c1_c3-d1_c6-h1_d2-e1_e4-f1_f6-g2WURCSPDB2Glycan 1.1.0
[]{[(4+1)][b-D-GlcpNAc]{[(4+1)][b-D-GlcpNAc]{[(4+1)][b-D-Manp]{[(3+1)][a-D-Manp]{[(2+1)][b-D-GlcpNAc]{[(4+1)][b-D-Galp]{[(6+2)][a-D-Neup5Ac]{}}}}[(6+1)][a-D-Manp]{}}}[(6+1)][a-L-Fucp]{}}}LINUCSPDB-CARE
#4: Chemical ChemComp-GOL / GLYCEROL / GLYCERIN / PROPANE-1,2,3-TRIOL


Mass: 92.094 Da / Num. of mol.: 3 / Source method: obtained synthetically / Formula: C3H8O3
#5: Water ChemComp-HOH / water


Mass: 18.015 Da / Num. of mol.: 13 / Source method: isolated from a natural source / Formula: H2O

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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: 3.56 Å3/Da / Density % sol: 65.45 %
Crystal growTemperature: 293 K / Method: vapor diffusion, sitting drop / pH: 7.5
Details: 0.1 M HEPES, pH 7.5, 30% v/v PEG400, 5% w/v PEG3000, 10% v/v glycerol, VAPOR DIFFUSION, SITTING DROP, temperature 293K

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Data collection

DiffractionMean temperature: 100 K
Diffraction sourceSource: SYNCHROTRON / Site: SSRL / Beamline: BL11-1 / Wavelength: 0.979 Å
DetectorType: PSI PILATUS 6M / Detector: PIXEL / Date: Jan 18, 2011
RadiationMonochromator: Side scattering bent cube-root I-beam single crystal, asymmetric cut 4.965 degrees
Protocol: SINGLE WAVELENGTH / Monochromatic (M) / Laue (L): M / Scattering type: x-ray
Radiation wavelengthWavelength: 0.979 Å / Relative weight: 1
ReflectionResolution: 2.8→40 Å / Num. all: 17491 / Num. obs: 17409 / % possible obs: 99.5 % / Observed criterion σ(F): 0 / Observed criterion σ(I): 0 / Redundancy: 7.2 % / Rsym value: 0.088 / Net I/σ(I): 19.8
Reflection shellResolution: 2.8→2.9 Å / Redundancy: 7.4 % / Mean I/σ(I) obs: 3.5 / Rsym value: 0.484 / % possible all: 99.9

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Processing

Software
NameVersionClassification
Blu-Icedata collection
PHASERphasing
PHENIX(phenix.refine: 1.7.2_869)refinement
XDSdata reduction
XDSdata scaling
RefinementMethod to determine structure: MOLECULAR REPLACEMENT
Starting model: PDB ENTRY 4JY6

4jy6


Resolution: 2.8→19.915 Å / SU ML: 0.76 / σ(F): 0 / Phase error: 21.65 / Stereochemistry target values: ML
RfactorNum. reflection% reflection
Rfree0.2341 870 5.01 %
Rwork0.1894 --
obs0.1917 17353 99.89 %
all-17491 -
Solvent computationShrinkage radii: 0.73 Å / VDW probe radii: 1 Å / Solvent model: FLAT BULK SOLVENT MODEL / Bsol: 20.37 Å2 / ksol: 0.357 e/Å3
Displacement parameters
Baniso -1Baniso -2Baniso -3
1--3.3525 Å20 Å2-0 Å2
2--10.1945 Å2-0 Å2
3----6.842 Å2
Refinement stepCycle: LAST / Resolution: 2.8→19.915 Å
ProteinNucleic acidLigandSolventTotal
Num. atoms3319 0 134 13 3466
Refine LS restraints
Refine-IDTypeDev idealNumber
X-RAY DIFFRACTIONf_bond_d0.013550
X-RAY DIFFRACTIONf_angle_d1.4434851
X-RAY DIFFRACTIONf_dihedral_angle_d23.9281299
X-RAY DIFFRACTIONf_chiral_restr0.201571
X-RAY DIFFRACTIONf_plane_restr0.006597
LS refinement shell
Resolution (Å)Rfactor RfreeNum. reflection RfreeRfactor RworkNum. reflection RworkRefine-ID% reflection obs (%)
2.8-2.97490.34951480.27272681X-RAY DIFFRACTION100
2.9749-3.20370.26651510.23412674X-RAY DIFFRACTION100
3.2037-3.52450.28951360.20112726X-RAY DIFFRACTION100
3.5245-4.03080.24051390.1962748X-RAY DIFFRACTION100
4.0308-5.06460.1911550.14972760X-RAY DIFFRACTION100
5.0646-19.91510.2031410.17542894X-RAY DIFFRACTION100
Refinement TLS params.

Method: refined / Refine-ID: X-RAY DIFFRACTION

IDL11 (°2)L12 (°2)L13 (°2)L22 (°2)L23 (°2)L33 (°2)S11 (Å °)S12 (Å °)S13 (Å °)S21 (Å °)S22 (Å °)S23 (Å °)S31 (Å °)S32 (Å °)S33 (Å °)T11 (Å2)T12 (Å2)T13 (Å2)T22 (Å2)T23 (Å2)T33 (Å2)Origin x (Å)Origin y (Å)Origin z (Å)
1-0.0718-0.19520.2492-0.0405-0.11110.33220.06250.2470.1505-0.3364-0.0347-0.009-0.12350.545400.51740.0849-0.01750.60810.02250.592522.467327.05730.9696
21.1238-0.3258-1.23130.4456-0.44991.1404-0.04050.2010.06480.28340.0028-0.09020.19030.231700.4520.0682-0.12870.3284-0.02910.338715.912224.908839.5005
3-0.01960.07030.14650.2942-0.2105-0.0375-0.0030.03450.06580.05110.2133-0.1883-0.11860.443700.2262-0.0051-0.02390.3347-0.04630.251813.615325.738824.1858
40.3328-0.5244-0.1608-0.179-0.09690.3426-0.0397-0.1547-0.0589-0.14990.14340.0195-0.0849-0.034200.1795-0.0028-0.00690.2647-0.01850.28616.529922.56361.1652
51.16431.25170.10250.37050.53360.5874-0.0913-0.23130.260.0842-0.09270.0415-0.10540.049300.27660.0688-0.05550.4158-0.05850.38336.930724.90273.9451
60.0780.0274-0.082-0.0605-0.22860.1292-0.02090.20040.31370.21430.02520.04320.03340.0339-00.2735-0.03050.00620.4128-0.00580.386814.010226.4714-2.3132
70.7828-0.0643-0.29480.62690.48610.73770.06490.20560.01780.2443-0.05030.26140.21550.0199-00.4368-0.00970.03620.2259-0.01670.3137-7.107619.771136.2569
80.05660.14480.05020.23780.23590.0443-0.9796-0.13470.7467-0.2520.8183-0.1325-0.01530.065700.5263-0.14670.01170.2660.07490.33840.060629.297434.2099
91.13110.1851-0.4170.39160.28870.84090.0125-0.0468-0.01690.2946-0.04420.16710.1037-0.1522-00.46230.00940.00250.1689-0.01090.2819-2.325928.250243.3963
100.28770.3872-0.19660.24220.0710.1148-0.07610.2872-0.10350.0792-0.14930.22090.04060.0119-00.39650.0202-0.10460.3331-0.01590.392-8.571420.946718.9714
111.8860.57350.32220.49580.22010.4416-0.0996-0.1509-0.14840.00490.0724-0.17670.00040.0162-00.23940.02740.02880.39160.0310.32731.354911.09432.1968
120.10460.06350.10660.56090.03260.0590.03130.456-0.69040.0358-0.15640.01760.0845-0.524500.2412-0.0262-0.02810.5165-0.04490.4466-6.28.2031-2.6826
Refinement TLS group
IDRefine-IDRefine TLS-IDSelection details
1X-RAY DIFFRACTION1chain 'A' and (resseq 9:26)
2X-RAY DIFFRACTION2chain 'A' and (resseq 27:92)
3X-RAY DIFFRACTION3chain 'A' and (resseq 93:119)
4X-RAY DIFFRACTION4chain 'A' and (resseq 120:151)
5X-RAY DIFFRACTION5chain 'A' and (resseq 152:188)
6X-RAY DIFFRACTION6chain 'A' and (resseq 189:211)
7X-RAY DIFFRACTION7chain 'B' and (resseq 1:39)
8X-RAY DIFFRACTION8chain 'B' and (resseq 40:51)
9X-RAY DIFFRACTION9chain 'B' and (resseq 52:101)
10X-RAY DIFFRACTION10chain 'B' and (resseq 102:117)
11X-RAY DIFFRACTION11chain 'B' and (resseq 118:201)
12X-RAY DIFFRACTION12chain 'B' and (resseq 202:211)

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