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- PDB-5j11: Structure of human TSLP in complex with TSLPR and IL-7Ralpha -

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Basic information

Entry
Database: PDB / ID: 5j11
TitleStructure of human TSLP in complex with TSLPR and IL-7Ralpha
Components
  • Cytokine receptor-like factor 2
  • Interleukin-7 receptor subunit alpha
  • Thymic stromal lymphopoietin
KeywordsSIGNALING PROTEIN / cytokine inflammation TSLP signaling complex
Function / homology
Function and homology information


positive regulation of chemokine (C-C motif) ligand 1 production / interleukin-7 receptor binding / interleukin-7 receptor activity / positive regulation of granulocyte colony-stimulating factor production / positive regulation of mast cell activation / negative regulation of T cell mediated cytotoxicity / regulation of DNA recombination / positive regulation of T cell differentiation in thymus / positive regulation of cytokine-mediated signaling pathway / positive regulation of receptor signaling pathway via STAT ...positive regulation of chemokine (C-C motif) ligand 1 production / interleukin-7 receptor binding / interleukin-7 receptor activity / positive regulation of granulocyte colony-stimulating factor production / positive regulation of mast cell activation / negative regulation of T cell mediated cytotoxicity / regulation of DNA recombination / positive regulation of T cell differentiation in thymus / positive regulation of cytokine-mediated signaling pathway / positive regulation of receptor signaling pathway via STAT / interleukin-7-mediated signaling pathway / positive regulation of interleukin-5 production / positive regulation of interleukin-13 production / negative regulation of T cell apoptotic process / cellular homeostasis / cytokine receptor activity / cytokine binding / regulation of cell size / T cell homeostasis / B cell homeostasis / hemopoiesis / B cell proliferation / positive regulation of interleukin-10 production / cell surface receptor signaling pathway via JAK-STAT / lymph node development / defense response to fungus / antigen binding / coreceptor activity / positive regulation of chemokine production / Interleukin-7 signaling / cytokine activity / positive regulation of receptor signaling pathway via JAK-STAT / T cell mediated cytotoxicity / clathrin-coated endocytic vesicle membrane / positive regulation of interleukin-6 production / cell morphogenesis / cytokine-mediated signaling pathway / positive regulation of inflammatory response / T cell differentiation in thymus / Cargo recognition for clathrin-mediated endocytosis / antimicrobial humoral immune response mediated by antimicrobial peptide / Clathrin-mediated endocytosis / gene expression / defense response to Gram-negative bacterium / cell surface receptor signaling pathway / receptor complex / defense response to Gram-positive bacterium / immune response / external side of plasma membrane / positive regulation of cell population proliferation / positive regulation of gene expression / negative regulation of apoptotic process / signal transduction / extracellular space / extracellular region / nucleoplasm / plasma membrane / cytosol
Similarity search - Function
Thymic stromal lymphopoietin / Immunoglobulin-like - #1870 / Thymic stromal lymphopoietin / Thymic stromal lymphopoietin superfamily / Thymic stromal lymphopoietin / IL-7Ralpha, fibronectin type III domain / Fibronectin type III domain / : / : / Cytokine receptor-like factor 2-like, D2 domain ...Thymic stromal lymphopoietin / Immunoglobulin-like - #1870 / Thymic stromal lymphopoietin / Thymic stromal lymphopoietin superfamily / Thymic stromal lymphopoietin / IL-7Ralpha, fibronectin type III domain / Fibronectin type III domain / : / : / Cytokine receptor-like factor 2-like, D2 domain / Cytokine receptor-like factor 2, domain 1 / Short hematopoietin receptor family 1 signature. / Short hematopoietin receptor, family 1, conserved site / Growth Hormone; Chain: A; / Fibronectin type III domain / Fibronectin type-III domain profile. / Fibronectin type III / Fibronectin type III superfamily / Immunoglobulin-like fold / Immunoglobulins / Up-down Bundle / Immunoglobulin-like / Sandwich / Mainly Beta / Mainly Alpha
Similarity search - Domain/homology
TRIETHYLENE GLYCOL / : / Interleukin-7 receptor subunit alpha / Thymic stromal lymphopoietin / Cytokine receptor-like factor 2
Similarity search - Component
Biological speciesHomo sapiens (human)
MethodX-RAY DIFFRACTION / SYNCHROTRON / MOLECULAR REPLACEMENT / Resolution: 2.56 Å
AuthorsVerstraete, K. / Savvides, S.N.
CitationJournal: Nat Commun / Year: 2017
Title: Structure and antagonism of the receptor complex mediated by human TSLP in allergy and asthma.
Authors: Kenneth Verstraete / Frank Peelman / Harald Braun / Juan Lopez / Dries Van Rompaey / Ann Dansercoer / Isabel Vandenberghe / Kris Pauwels / Jan Tavernier / Bart N Lambrecht / Hamida Hammad / ...Authors: Kenneth Verstraete / Frank Peelman / Harald Braun / Juan Lopez / Dries Van Rompaey / Ann Dansercoer / Isabel Vandenberghe / Kris Pauwels / Jan Tavernier / Bart N Lambrecht / Hamida Hammad / Hans De Winter / Rudi Beyaert / Guy Lippens / Savvas N Savvides /
Abstract: The pro-inflammatory cytokine thymic stromal lymphopoietin (TSLP) is pivotal to the pathophysiology of widespread allergic diseases mediated by type 2 helper T cell (Th2) responses, including asthma ...The pro-inflammatory cytokine thymic stromal lymphopoietin (TSLP) is pivotal to the pathophysiology of widespread allergic diseases mediated by type 2 helper T cell (Th2) responses, including asthma and atopic dermatitis. The emergence of human TSLP as a clinical target against asthma calls for maximally harnessing its therapeutic potential via structural and mechanistic considerations. Here we employ an integrative experimental approach focusing on productive and antagonized TSLP complexes and free cytokine. We reveal how cognate receptor TSLPR allosterically activates TSLP to potentiate the recruitment of the shared interleukin 7 receptor α-chain (IL-7Rα) by leveraging the flexibility, conformational heterogeneity and electrostatics of the cytokine. We further show that the monoclonal antibody Tezepelumab partly exploits these principles to neutralize TSLP activity. Finally, we introduce a fusion protein comprising a tandem of the TSLPR and IL-7Rα extracellular domains, which harnesses the mechanistic intricacies of the TSLP-driven receptor complex to manifest high antagonistic potency.
History
DepositionMar 28, 2016Deposition site: RCSB / Processing site: PDBE
Revision 1.0Apr 5, 2017Provider: repository / Type: Initial release
Revision 1.1Apr 19, 2017Group: Database references
Revision 1.2Feb 28, 2018Group: Source and taxonomy / Category: entity_src_gen
Item: _entity_src_gen.host_org_common_name / _entity_src_gen.pdbx_host_org_strain / _entity_src_gen.pdbx_host_org_variant
Revision 1.3Jul 29, 2020Group: Data collection / Derived calculations / Structure summary
Category: chem_comp / entity ...chem_comp / entity / pdbx_chem_comp_identifier / pdbx_entity_nonpoly / struct_conn / struct_site / struct_site_gen
Item: _chem_comp.name / _chem_comp.type ..._chem_comp.name / _chem_comp.type / _entity.pdbx_description / _pdbx_entity_nonpoly.name / _struct_conn.pdbx_role
Description: Carbohydrate remediation / Provider: repository / Type: Remediation
Revision 1.4Jan 10, 2024Group: 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
Revision 1.5Nov 20, 2024Group: Structure summary / Category: pdbx_entry_details / pdbx_modification_feature

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

Downloads & links

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Assembly

Deposited unit
A: Thymic stromal lymphopoietin
B: Interleukin-7 receptor subunit alpha
C: Cytokine receptor-like factor 2
hetero molecules


Theoretical massNumber of molelcules
Total (without water)71,2826
Polymers70,6903
Non-polymers5933
Water52229
1


  • Idetical with deposited unit
  • defined by software
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1
Buried area4660 Å2
ΔGint1 kcal/mol
Surface area24620 Å2
MethodPISA
Unit cell
Length a, b, c (Å)135.783, 66.644, 91.971
Angle α, β, γ (deg.)90.00, 109.20, 90.00
Int Tables number5
Space group name H-MC121

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Components

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Protein , 3 types, 3 molecules ABC

#1: Protein Thymic stromal lymphopoietin


Mass: 16523.906 Da / Num. of mol.: 1
Mutation: Residues 127 to 131 were deleted in the construct used for crystallisation.
Source method: isolated from a genetically manipulated source
Details: Before crystallisation, the N-terminal His-tag (residues 1 - 17, MGSSHHHHHHSSGLVPR) was removed by thrombin cleavage. Residues 127 to 131 of TSLP (127-RRKRK-131) (according to the reference ...Details: Before crystallisation, the N-terminal His-tag (residues 1 - 17, MGSSHHHHHHSSGLVPR) was removed by thrombin cleavage. Residues 127 to 131 of TSLP (127-RRKRK-131) (according to the reference sequence numbering scheme for TSLP) were deleted in the construct used for crystallization.
Source: (gene. exp.) Homo sapiens (human) / Gene: TSLP / Plasmid: pET15b-hTSLPdelta127-131
Details (production host): ORF cloned between NdeI and BamHI sites
Production host: Escherichia coli BL21(DE3) (bacteria) / References: UniProt: Q969D9
#2: Protein Interleukin-7 receptor subunit alpha


Mass: 27592.170 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Details: Before crystallisation, the N-terminal His-tag (residues 1 - 17, MGSSHHHHHHSSGLVPR) was removed by thrombin cleavage.
Source: (gene. exp.) Homo sapiens (human) / Gene: IL7R / Plasmid: pET15b-hIL7Ralpha
Details (production host): ORF cloned between NdeI and BamHI sites
Production host: Escherichia coli BL21(DE3) (bacteria) / References: UniProt: D6RGV2, UniProt: P16871*PLUS
#3: Protein Cytokine receptor-like factor 2 / Cytokine receptor-like 2 / IL-XR / Thymic stromal lymphopoietin protein receptor / TSLP receptor


Mass: 26573.562 Da / Num. of mol.: 1 / Mutation: N47Q
Source method: isolated from a genetically manipulated source
Details: The signal peptide (residues 1 - 24) is removed from the mature protein.
Source: (gene. exp.) Homo sapiens (human) / Gene: CRLF2, CRL2, ILXR, TSLPR / Plasmid: pcDNA4/TO-hTSLPR-N47Q-His
Details (production host): stable cell line, tetracycline inducible expression
Cell line (production host): HEK293S MGAT1-/- / Production host: Homo sapiens (human) / References: UniProt: Q9HC73

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Sugars , 1 types, 2 molecules

#5: 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


Type: D-saccharide, beta linking / Mass: 221.208 Da / Num. of mol.: 2
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 , 2 types, 30 molecules

#4: Chemical ChemComp-PGE / TRIETHYLENE GLYCOL


Mass: 150.173 Da / Num. of mol.: 1 / Source method: obtained synthetically / Formula: C6H14O4
#6: Water ChemComp-HOH / water


Mass: 18.015 Da / Num. of mol.: 29 / Source method: isolated from a natural source / Formula: H2O

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Details

Has protein modificationY

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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: 3.05 Å3/Da / Density % sol: 59.72 %
Crystal growTemperature: 293 K / Method: vapor diffusion, sitting drop / pH: 8.8
Details: 0.02 M Citric acid 0.08 M BIS-TRIS propane 16% w/v polyethylene glycol 3350

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Data collection

DiffractionMean temperature: 100 K
Diffraction sourceSource: SYNCHROTRON / Site: SOLEIL / Beamline: PROXIMA 2 / Wavelength: 0.9801 Å
DetectorType: ADSC QUANTUM 315r / Detector: CCD / Date: Jan 30, 2015
RadiationMonochromator: Si[111] / Protocol: SINGLE WAVELENGTH / Monochromatic (M) / Laue (L): M / Scattering type: x-ray
Radiation wavelengthWavelength: 0.9801 Å / Relative weight: 1
ReflectionResolution: 2.56→50 Å / Num. obs: 24638 / % possible obs: 97.8 % / Redundancy: 3.2 % / Biso Wilson estimate: 69.9 Å2 / CC1/2: 0.998 / Rrim(I) all: 0.062 / Net I/σ(I): 14.2
Reflection shellResolution: 2.56→2.72 Å / Redundancy: 3.1 % / Mean I/σ(I) obs: 1.7 / Rrim(I) all: 0.776 / % possible all: 94.2

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Processing

Software
NameVersionClassification
BUSTER2.10.2refinement
XDSVERSION November 3, 2014data reduction
PHASERphasing
Coot0.8.1model building
RefinementMethod to determine structure: MOLECULAR REPLACEMENT
Starting model: X-ray structure of human TSLP in complex with human TSLPR and mouse IL-7Ralpha; PDB 3DI2: chain B

3di2


Resolution: 2.56→45.76 Å / Cor.coef. Fo:Fc: 0.9349 / Cor.coef. Fo:Fc free: 0.9285 / Cross valid method: THROUGHOUT / σ(F): 0 / SU R Blow DPI: 0.319 / SU Rfree Blow DPI: 0.219
RfactorNum. reflection% reflectionSelection details
Rfree0.2176 1238 5.02 %RANDOM
Rwork0.1912 ---
obs0.1926 24638 97.65 %-
Displacement parametersBiso mean: 88.73 Å2
Baniso -1Baniso -2Baniso -3
1-9.2551 Å20 Å212.6844 Å2
2---2.5336 Å20 Å2
3----6.7216 Å2
Refine analyzeLuzzati coordinate error obs: 0.328 Å
Refinement stepCycle: LAST / Resolution: 2.56→45.76 Å
ProteinNucleic acidLigandSolventTotal
Num. atoms3966 0 38 29 4033
Refine LS restraints
Refine-IDTypeDev idealNumberRestraint functionWeight
X-RAY DIFFRACTIONt_bond_d0.017863HARMONIC2
X-RAY DIFFRACTIONt_angle_deg1.0914104HARMONIC2
X-RAY DIFFRACTIONt_dihedral_angle_d1679SINUSOIDAL2
X-RAY DIFFRACTIONt_incorr_chiral_ct
X-RAY DIFFRACTIONt_pseud_angle
X-RAY DIFFRACTIONt_trig_c_planes92HARMONIC2
X-RAY DIFFRACTIONt_gen_planes1172HARMONIC5
X-RAY DIFFRACTIONt_it7863HARMONIC20
X-RAY DIFFRACTIONt_nbd
X-RAY DIFFRACTIONt_omega_torsion3.58
X-RAY DIFFRACTIONt_other_torsion15.73
X-RAY DIFFRACTIONt_improper_torsion
X-RAY DIFFRACTIONt_chiral_improper_torsion549SEMIHARMONIC5
X-RAY DIFFRACTIONt_sum_occupancies
X-RAY DIFFRACTIONt_utility_distance
X-RAY DIFFRACTIONt_utility_angle
X-RAY DIFFRACTIONt_utility_torsion
X-RAY DIFFRACTIONt_ideal_dist_contact7975SEMIHARMONIC4
LS refinement shellResolution: 2.56→2.67 Å / Total num. of bins used: 12
RfactorNum. reflection% reflection
Rfree0.3351 138 5.04 %
Rwork0.2674 2600 -
all0.2708 2738 -
obs--89.97 %
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)
14.44461.175-1.02645.6989-0.6781.7977-0.0655-0.0246-0.48460.025-0.1559-0.73210.03140.3730.2214-0.3384-0.04330.0083-0.0024-0.0301-0.0399290.263941.911336.0957
24.11530.554-0.50533.3084-1.14892.8863-0.1490.37050.2808-0.3638-0.569-1.1066-0.59981.01220.718-0.2394-0.2753-0.0120.08960.3058-0.0825303.27862.908719.9274
33.7777-1.44672.00612.2414-1.64712.9168-0.16760.10140.056-0.0009-0.03810.2181-0.81610.01380.20570.19460.0294-0.1174-0.161-0.0794-0.328274.252868.488813.6108
44.1416-0.3469-0.13195.5834-1.72092.67630.09990.186-0.87-0.4587-0.05890.23420.30640.0917-0.041-0.31620.0153-0.0137-0.1791-0.12330.0929267.4425.668539.5052
53.0353-2.53472.02326.8271-3.17024.2026-0.08510.09060.14710.12560.14650.6906-1.1017-0.4863-0.0614-0.09090.14860.0082-0.1282-0.1158-0.1312260.878756.328.2342
Refinement TLS group
IDRefine-IDRefine TLS-IDSelection details
1X-RAY DIFFRACTION1{ A|* }
2X-RAY DIFFRACTION2{ B|37 - B|124 }
3X-RAY DIFFRACTION3{ B|125 - B|231 }
4X-RAY DIFFRACTION4{ C|29 - C|114 C|500}
5X-RAY DIFFRACTION5{ C|115 - C|219 C|501}

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