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- PDB-4m5y: Crystal structure of broadly neutralizing Fab 5J8 -

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Basic information

Entry
Database: PDB / ID: 4m5y
TitleCrystal structure of broadly neutralizing Fab 5J8
Components
  • Fab 5J8 heavy chain
  • Fab 5J8 light chain
KeywordsIMMUNE SYSTEM / immunoglobulin fold / fragment antigen binding / influenza hemagglutinin receptor binding site
Function / homologyImmunoglobulins / Immunoglobulin-like / Sandwich / Mainly Beta / DI(HYDROXYETHYL)ETHER
Function and homology information
Biological speciesHomo sapiens (human)
MethodX-RAY DIFFRACTION / SYNCHROTRON / MOLECULAR REPLACEMENT / Resolution: 1.55 Å
AuthorsLee, P.S. / Wilson, I.A.
CitationJournal: J Virol / Year: 2013
Title: Antibody recognition of the pandemic H1N1 Influenza virus hemagglutinin receptor binding site.
Authors: Minsun Hong / Peter S Lee / Ryan M B Hoffman / Xueyong Zhu / Jens C Krause / Nick S Laursen / Sung-Il Yoon / Langzhou Song / Lynda Tussey / James E Crowe / Andrew B Ward / Ian A Wilson
Abstract: Influenza virus is a global health concern due to its unpredictable pandemic potential. This potential threat was realized in 2009 when an H1N1 virus emerged that resembled the 1918 virus in ...Influenza virus is a global health concern due to its unpredictable pandemic potential. This potential threat was realized in 2009 when an H1N1 virus emerged that resembled the 1918 virus in antigenicity but fortunately was not nearly as deadly. 5J8 is a human antibody that potently neutralizes a broad spectrum of H1N1 viruses, including the 1918 and 2009 pandemic viruses. Here, we present the crystal structure of 5J8 Fab in complex with a bacterially expressed and refolded globular head domain from the hemagglutinin (HA) of the A/California/07/2009 (H1N1) pandemic virus. 5J8 recognizes a conserved epitope in and around the receptor binding site (RBS), and its HCDR3 closely mimics interactions of the sialic acid receptor. Electron microscopy (EM) reconstructions of 5J8 Fab in complex with an HA trimer from a 1986 H1 strain and with an engineered stabilized HA trimer from the 2009 H1 pandemic virus showed a similar mode of binding. As for other characterized RBS-targeted antibodies, 5J8 uses avidity to extend its breadth and affinity against divergent H1 strains. 5J8 selectively interacts with HA insertion residue 133a, which is conserved in pandemic H1 strains and has precluded binding of other RBS-targeted antibodies. Thus, the RBS of divergent HAs is targeted by 5J8 and adds to the growing arsenal of common recognition motifs for design of therapeutics and vaccines. Moreover, consistent with previous studies, the bacterially expressed H1 HA properly refolds, retaining its antigenic structure, and presents a low-cost and rapid alternative for engineering and manufacturing candidate flu vaccines.
History
DepositionAug 8, 2013Deposition site: RCSB / Processing site: RCSB
Revision 1.0Sep 25, 2013Provider: repository / Type: Initial release
Revision 1.1Nov 6, 2013Group: Database references
Revision 1.2Sep 20, 2023Group: Data collection / Database references ...Data collection / Database references / Derived calculations / Refinement description
Category: chem_comp_atom / chem_comp_bond ...chem_comp_atom / chem_comp_bond / database_2 / pdbx_initial_refinement_model / struct_site
Item: _database_2.pdbx_DOI / _database_2.pdbx_database_accession ..._database_2.pdbx_DOI / _database_2.pdbx_database_accession / _struct_site.pdbx_auth_asym_id / _struct_site.pdbx_auth_comp_id / _struct_site.pdbx_auth_seq_id

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

Downloads & links

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Assembly

Deposited unit
H: Fab 5J8 heavy chain
L: Fab 5J8 light chain
I: Fab 5J8 heavy chain
M: Fab 5J8 light chain
hetero molecules


Theoretical massNumber of molelcules
Total (without water)97,16417
Polymers95,6964
Non-polymers1,46813
Water15,601866
1
H: Fab 5J8 heavy chain
L: Fab 5J8 light chain
hetero molecules


Theoretical massNumber of molelcules
Total (without water)48,5738
Polymers47,8482
Non-polymers7256
Water362
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1
Buried area5280 Å2
ΔGint-12 kcal/mol
Surface area19530 Å2
MethodPISA
2
I: Fab 5J8 heavy chain
M: Fab 5J8 light chain
hetero molecules


Theoretical massNumber of molelcules
Total (without water)48,5919
Polymers47,8482
Non-polymers7437
Water362
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1
Buried area5080 Å2
ΔGint-14 kcal/mol
Surface area20010 Å2
MethodPISA
Unit cell
Length a, b, c (Å)90.573, 99.983, 144.333
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 Fab 5J8 heavy chain


Mass: 25088.070 Da / Num. of mol.: 2
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Production host: Trichoplusia ni (cabbage looper)
#2: Antibody Fab 5J8 light chain


Mass: 22760.098 Da / Num. of mol.: 2
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Production host: Trichoplusia ni (cabbage looper)
#3: Chemical
ChemComp-PEG / DI(HYDROXYETHYL)ETHER


Mass: 106.120 Da / Num. of mol.: 12 / Source method: obtained synthetically / Formula: C4H10O3
#4: Chemical ChemComp-PG4 / TETRAETHYLENE GLYCOL


Mass: 194.226 Da / Num. of mol.: 1 / Source method: obtained synthetically / Formula: C8H18O5 / Comment: precipitant*YM
#5: Water ChemComp-HOH / water


Mass: 18.015 Da / Num. of mol.: 866 / 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: 3.41 Å3/Da / Density % sol: 63.98 %
Crystal growTemperature: 277 K / Method: vapor diffusion, sitting drop / pH: 8
Details: 0.2 M calcium acetate, 11% PEG3350, pH 8, VAPOR DIFFUSION, SITTING DROP, temperature 277K

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Data collection

DiffractionMean temperature: 77 K
Diffraction sourceSource: SYNCHROTRON / Site: APS / Beamline: 23-ID-D / Wavelength: 1.0332 Å
DetectorType: MARMOSAIC 300 mm CCD / Detector: CCD / Date: Nov 17, 2012
RadiationMonochromator: Double crystal cryo-cooled Si(111) / Protocol: SINGLE WAVELENGTH / Monochromatic (M) / Laue (L): M / Scattering type: x-ray
Radiation wavelengthWavelength: 1.0332 Å / Relative weight: 1
ReflectionResolution: 1.55→50 Å / Num. all: 190412 / Num. obs: 183997 / % possible obs: 96.9 % / Observed criterion σ(I): -3 / Redundancy: 7.3 % / Biso Wilson estimate: 20.1 Å2 / Rmerge(I) obs: 0.046 / Rsym value: 0.046 / Net I/σ(I): 21.9
Reflection shellResolution: 1.55→1.57 Å / Redundancy: 5.6 % / Rmerge(I) obs: 0.91 / Mean I/σ(I) obs: 1.8 / Num. unique all: 6687 / Rsym value: 0.91 / % possible all: 72.5

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Processing

Software
NameVersionClassification
Blu-Icedata collection
PHASERphasing
PHENIX(phenix.refine: 1.8.1_1168)refinement
XDSdata reduction
XDSdata scaling
RefinementMethod to determine structure: MOLECULAR REPLACEMENT
Starting model: PDB ENTRY 3MLY CHAINS H AND L

3mly


Resolution: 1.55→49.151 Å / SU ML: 0.13 / σ(F): 2 / Phase error: 19.02 / Stereochemistry target values: ML
RfactorNum. reflection% reflectionSelection details
Rfree0.1865 9196 5.01 %RANDOM
Rwork0.1723 ---
obs0.173 183666 96.81 %-
Solvent computationShrinkage radii: 0.9 Å / VDW probe radii: 1.11 Å / Solvent model: FLAT BULK SOLVENT MODEL
Refinement stepCycle: LAST / Resolution: 1.55→49.151 Å
ProteinNucleic acidLigandSolventTotal
Num. atoms6494 0 97 866 7457
Refine LS restraints
Refine-IDTypeDev idealNumber
X-RAY DIFFRACTIONf_bond_d0.0096900
X-RAY DIFFRACTIONf_angle_d1.2639437
X-RAY DIFFRACTIONf_dihedral_angle_d14.1662495
X-RAY DIFFRACTIONf_chiral_restr0.0851082
X-RAY DIFFRACTIONf_plane_restr0.0071193
LS refinement shell
Resolution (Å)Rfactor RfreeNum. reflection RfreeRfactor RworkNum. reflection RworkRefine-ID% reflection obs (%)
1.55-1.56760.29352380.26824353X-RAY DIFFRACTION73
1.5676-1.58610.25282540.24784617X-RAY DIFFRACTION78
1.5861-1.60540.24562410.24125029X-RAY DIFFRACTION85
1.6054-1.62570.26382600.24195539X-RAY DIFFRACTION93
1.6257-1.64710.21883330.2155792X-RAY DIFFRACTION98
1.6471-1.66970.22073250.20385821X-RAY DIFFRACTION98
1.6697-1.69360.19913090.18965854X-RAY DIFFRACTION98
1.6936-1.71880.20593560.18415822X-RAY DIFFRACTION98
1.7188-1.74570.19143070.17845863X-RAY DIFFRACTION99
1.7457-1.77430.21882950.18475919X-RAY DIFFRACTION99
1.7743-1.80490.20943340.18155810X-RAY DIFFRACTION99
1.8049-1.83770.19562920.17215918X-RAY DIFFRACTION99
1.8377-1.87310.18323260.16585922X-RAY DIFFRACTION99
1.8731-1.91130.1913130.16395904X-RAY DIFFRACTION99
1.9113-1.95290.1753160.16675902X-RAY DIFFRACTION99
1.9529-1.99830.18633280.1715926X-RAY DIFFRACTION99
1.9983-2.04830.20353240.17155935X-RAY DIFFRACTION99
2.0483-2.10370.18173020.16465940X-RAY DIFFRACTION99
2.1037-2.16560.19083070.16145954X-RAY DIFFRACTION99
2.1656-2.23550.17483040.16555958X-RAY DIFFRACTION99
2.2355-2.31540.19163270.17085982X-RAY DIFFRACTION100
2.3154-2.40810.18872930.17146032X-RAY DIFFRACTION100
2.4081-2.51760.19353210.17865983X-RAY DIFFRACTION100
2.5176-2.65040.19643230.18086010X-RAY DIFFRACTION100
2.6504-2.81640.19742930.17856037X-RAY DIFFRACTION100
2.8164-3.03380.19373310.18156050X-RAY DIFFRACTION100
3.0338-3.33910.19223090.1836093X-RAY DIFFRACTION100
3.3391-3.82210.17923130.17456105X-RAY DIFFRACTION100
3.8221-4.81480.14982980.14086172X-RAY DIFFRACTION100
4.8148-49.17520.16683240.15816228X-RAY DIFFRACTION98
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)
10.9358-0.0444-0.93920.54030.42972.7555-0.05430.02310.02730.05780.0378-0.00930.08670.03510.01980.08260.0106-0.00710.09260.00510.116835.97115.068817.0715
21.16630.5326-0.28811.84590.21721.1998-0.1698-0.4579-0.7533-0.2296-0.2782-0.91650.31020.46410.28740.2820.08010.18230.34450.24460.661233.5976-4.170243.461
31.36191.0951-0.27971.89030.25072.5433-0.11080.1799-0.148-0.0069-0.02210.0950.6153-0.33070.05620.2354-0.09370.05010.2026-0.04660.160224.907-0.25886.5246
42.06081.2463-0.88612.5878-0.60892.9801-0.2158-0.0582-0.2811-0.58990.0107-0.34220.14030.11530.09530.2634-0.03030.07470.15550.03470.186717.3828-9.331843.6578
50.85240.0409-0.74180.4406-0.28952.8973-0.0542-0.05340.0417-0.04640.0336-0.00030.05930.00290.02360.0837-0.00850.00030.0972-0.01150.136954.597115.1167-14.2791
61.2998-0.14930.20583.7330.38092.0675-0.01690.0353-0.28660.1526-0.11290.82450.2873-0.17790.0650.19790.00870.08580.1634-0.03570.364657.8786-6.0149-40.1282
71.2147-1.0709-0.21581.61130.04152.8445-0.0672-0.1639-0.06290.0655-0.0124-0.13670.53890.45420.02650.2080.10290.02650.25260.02690.16366.36310.4093-3.6137
81.3159-0.5943-0.42941.83270.36414.0362-0.1815-0.1563-0.07010.3290.17940.0680.11220.1833-0.01170.17940.09330.02540.1859-0.00290.146373.5848-9.5158-40.5857
Refinement TLS group
IDRefine-IDRefine TLS-IDSelection detailsAuth asym-IDAuth seq-ID
1X-RAY DIFFRACTION1chain H and resid 1:123H1 - 123
2X-RAY DIFFRACTION2chain H and resid 124:209H124 - 209
3X-RAY DIFFRACTION3chain L and resid 1:109L1 - 109
4X-RAY DIFFRACTION4chain L and resid 110:209L110 - 209
5X-RAY DIFFRACTION5chain I and resid 1:123I1 - 123
6X-RAY DIFFRACTION6chain I and resid 124:209I124 - 209
7X-RAY DIFFRACTION7chain M and resid 1:109M1 - 109
8X-RAY DIFFRACTION8chain M and resid 110:209M110 - 209

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