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- PDB-5wb2: US28 bound to engineered chemokine CX3CL1.35 and nanobodies -

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Basic information

Entry
Database: PDB / ID: 5wb2
TitleUS28 bound to engineered chemokine CX3CL1.35 and nanobodies
Components
  • CX3CL1 protein
  • Envelope protein US28, nanobody 7 fusion protein
  • nanobody B1
KeywordsMEMBRANE PROTEIN / chemokine receptor / engineered proteins
Function / homology
Function and homology information


: / CXCR1 chemokine receptor binding / positive regulation of calcium-independent cell-cell adhesion / negative regulation of glutamate receptor signaling pathway / negative regulation of interleukin-1 alpha production / leukocyte adhesive activation / CX3C chemokine receptor binding / negative regulation of hippocampal neuron apoptotic process / autocrine signaling / synapse pruning ...: / CXCR1 chemokine receptor binding / positive regulation of calcium-independent cell-cell adhesion / negative regulation of glutamate receptor signaling pathway / negative regulation of interleukin-1 alpha production / leukocyte adhesive activation / CX3C chemokine receptor binding / negative regulation of hippocampal neuron apoptotic process / autocrine signaling / synapse pruning / negative regulation of neuron migration / positive regulation of microglial cell migration / negative regulation of microglial cell activation / positive regulation of transforming growth factor beta1 production / regulation of lipopolysaccharide-mediated signaling pathway / chemokine receptor activity / positive regulation of I-kappaB phosphorylation / microglial cell proliferation / positive regulation of actin filament bundle assembly / CCR chemokine receptor binding / leukocyte migration involved in inflammatory response / lymphocyte chemotaxis / integrin activation / eosinophil chemotaxis / angiogenesis involved in wound healing / chemokine-mediated signaling pathway / leukocyte chemotaxis / Chemokine receptors bind chemokines / chemokine activity / negative regulation of cell-substrate adhesion / positive regulation of cell-matrix adhesion / negative regulation of interleukin-1 beta production / neuron remodeling / positive regulation of neuroblast proliferation / positive chemotaxis / chemoattractant activity / monocyte chemotaxis / negative regulation of interleukin-6 production / negative regulation of apoptotic signaling pathway / negative regulation of tumor necrosis factor production / regulation of neurogenesis / negative regulation of extrinsic apoptotic signaling pathway in absence of ligand / cellular response to interleukin-1 / extrinsic apoptotic signaling pathway in absence of ligand / virus-mediated perturbation of host defense response / cell chemotaxis / neutrophil chemotaxis / negative regulation of cell migration / positive regulation of release of sequestered calcium ion into cytosol / cell projection / response to ischemia / microglial cell activation / positive regulation of smooth muscle cell proliferation / regulation of synaptic plasticity / defense response / positive regulation of neuron projection development / cell-cell adhesion / positive regulation of inflammatory response / cellular response to type II interferon / neuron cellular homeostasis / cytokine-mediated signaling pathway / chemotaxis / integrin binding / cell-cell signaling / cellular response to tumor necrosis factor / G alpha (i) signalling events / positive regulation of canonical NF-kappaB signal transduction / positive regulation of MAPK cascade / positive regulation of ERK1 and ERK2 cascade / positive regulation of phosphatidylinositol 3-kinase/protein kinase B signal transduction / cell adhesion / neuron projection / immune response / inflammatory response / G protein-coupled receptor signaling pathway / signaling receptor binding / neuronal cell body / viral envelope / positive regulation of cell population proliferation / negative regulation of apoptotic process / host cell plasma membrane / perinuclear region of cytoplasm / cell surface / positive regulation of transcription by RNA polymerase II / extracellular space / extracellular region / membrane / plasma membrane
Similarity search - Function
CX3C chemokine domain / Chemokine receptor family / Chemokine beta/gamma/delta / Intercrine alpha family (small cytokine C-X-C) (chemokine CXC). / Chemokine interleukin-8-like domain / Chemokine interleukin-8-like superfamily / Small cytokines (intecrine/chemokine), interleukin-8 like / OB fold (Dihydrolipoamide Acetyltransferase, E2P) - #40 / Rhopdopsin 7-helix transmembrane proteins / Rhodopsin 7-helix transmembrane proteins ...CX3C chemokine domain / Chemokine receptor family / Chemokine beta/gamma/delta / Intercrine alpha family (small cytokine C-X-C) (chemokine CXC). / Chemokine interleukin-8-like domain / Chemokine interleukin-8-like superfamily / Small cytokines (intecrine/chemokine), interleukin-8 like / OB fold (Dihydrolipoamide Acetyltransferase, E2P) - #40 / Rhopdopsin 7-helix transmembrane proteins / Rhodopsin 7-helix transmembrane proteins / G-protein coupled receptors family 1 signature. / G protein-coupled receptor, rhodopsin-like / GPCR, rhodopsin-like, 7TM / G-protein coupled receptors family 1 profile. / 7 transmembrane receptor (rhodopsin family) / OB fold (Dihydrolipoamide Acetyltransferase, E2P) / Up-down Bundle / Beta Barrel / Mainly Beta / Mainly Alpha
Similarity search - Domain/homology
CHOLESTEROL / (2R)-2,3-dihydroxypropyl (9Z)-octadec-9-enoate / G-protein coupled receptor homolog US28 / Fractalkine / CX3CL1 protein / Envelope protein US28
Similarity search - Component
Biological speciesHuman cytomegalovirus
Lama glama (llama)
Homo sapiens (human)
synthetic construct (others)
MethodX-RAY DIFFRACTION / SYNCHROTRON / MOLECULAR REPLACEMENT / Resolution: 3.5 Å
AuthorsJude, K.M. / Burg, J.S. / Tsutsumi, N. / Miles, T.F. / Garcia, K.C.
Funding support United States, 1items
OrganizationGrant numberCountry
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)GM097015 United States
CitationJournal: Elife / Year: 2018
Title: Viral GPCR US28 can signal in response to chemokine agonists of nearly unlimited structural degeneracy.
Authors: Miles, T.F. / Spiess, K. / Jude, K.M. / Tsutsumi, N. / Burg, J.S. / Ingram, J.R. / Waghray, D. / Hjorto, G.M. / Larsen, O. / Ploegh, H.L. / Rosenkilde, M.M. / Garcia, K.C.
History
DepositionJun 27, 2017Deposition site: RCSB / Processing site: RCSB
Revision 1.0Jun 13, 2018Provider: repository / Type: Initial release
Revision 1.1Dec 26, 2018Group: Data collection / Database references / Category: citation / citation_author
Item: _citation.country / _citation.journal_abbrev ..._citation.country / _citation.journal_abbrev / _citation.journal_id_CSD / _citation.journal_id_ISSN / _citation.journal_volume / _citation.pdbx_database_id_DOI / _citation.pdbx_database_id_PubMed / _citation.title / _citation.year
Revision 2.0Jan 1, 2020Group: Author supporting evidence / Polymer sequence / Category: entity_poly / pdbx_audit_support
Item: _entity_poly.pdbx_seq_one_letter_code_can / _pdbx_audit_support.funding_organization
Revision 2.1Oct 4, 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_conn
Item: _database_2.pdbx_DOI / _database_2.pdbx_database_accession ..._database_2.pdbx_DOI / _database_2.pdbx_database_accession / _struct_conn.conn_type_id / _struct_conn.id / _struct_conn.pdbx_dist_value / _struct_conn.pdbx_leaving_atom_flag / _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.ptnr1_symmetry / _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_atom_id / _struct_conn.ptnr2_label_comp_id / _struct_conn.ptnr2_label_seq_id / _struct_conn.ptnr2_symmetry

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

Downloads & links

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Assembly

Deposited unit
A: Envelope protein US28, nanobody 7 fusion protein
B: CX3CL1 protein
D: nanobody B1
hetero molecules


Theoretical massNumber of molelcules
Total (without water)77,40111
Polymers74,9413
Non-polymers2,4608
Water0
1


  • Idetical with deposited unit
  • defined by author&software
  • Evidence: gel filtration
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1
Buried area6620 Å2
ΔGint-45 kcal/mol
Surface area28810 Å2
MethodPISA
Unit cell
Length a, b, c (Å)47.170, 128.940, 127.740
Angle α, β, γ (deg.)90.00, 90.00, 90.00
Int Tables number18
Space group name H-MP21212

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Components

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Protein , 1 types, 1 molecules B

#2: Protein CX3CL1 protein / / engineered chemokine CX3CL1.35


Mass: 8631.051 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: CX3CL1 / Production host: Trichoplusia ni (cabbage looper) / References: UniProt: Q6I9S9, UniProt: P78423*PLUS

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Antibody , 2 types, 2 molecules AD

#1: Antibody Envelope protein US28, nanobody 7 fusion protein / / US28 protein


Mass: 52158.414 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Human cytomegalovirus, (gene. exp.) Lama glama (llama)
Gene: US28 / Cell line (production host): HEK293F / Production host: Homo sapiens (human) / References: UniProt: Q80KM9, UniProt: P69333*PLUS
#3: Antibody nanobody B1


Mass: 14151.589 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) synthetic construct (others) / Production host: Escherichia coli (E. coli)

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Non-polymers , 4 types, 8 molecules

#4: Chemical ChemComp-CLR / CHOLESTEROL / Cholesterol


Mass: 386.654 Da / Num. of mol.: 2 / Source method: obtained synthetically / Formula: C27H46O
#5: Chemical
ChemComp-OLC / (2R)-2,3-dihydroxypropyl (9Z)-octadec-9-enoate / 1-Oleoyl-R-glycerol


Mass: 356.540 Da / Num. of mol.: 4 / Source method: obtained synthetically / Formula: C21H40O4
#6: Chemical ChemComp-ZN / ZINC ION


Mass: 65.409 Da / Num. of mol.: 1 / Source method: obtained synthetically / Formula: Zn
#7: Chemical ChemComp-MES / 2-(N-MORPHOLINO)-ETHANESULFONIC ACID / MES (buffer)


Mass: 195.237 Da / Num. of mol.: 1 / Source method: obtained synthetically / Formula: C6H13NO4S / Comment: pH buffer*YM

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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: 2.59 Å3/Da / Density % sol: 52.54 %
Crystal growTemperature: 293 K / Method: lipidic cubic phase / pH: 6
Details: MES pH 6.0, 50 mM lithium sulfate, 35-39% PEG300, 1% 1,2,3-heptanetriol in monoolein:cholesterol (9:1)

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Data collection

DiffractionMean temperature: 100 K
Diffraction sourceSource: SYNCHROTRON / Site: APS / Beamline: 23-ID-D / Wavelength: 1.03323 Å
DetectorType: DECTRIS PILATUS3 S 6M / Detector: PIXEL / Date: Aug 7, 2016
RadiationMonochromator: double crystal Si(111) / Protocol: SINGLE WAVELENGTH / Monochromatic (M) / Laue (L): M / Scattering type: x-ray
Radiation wavelengthWavelength: 1.03323 Å / Relative weight: 1
ReflectionResolution: 3.5→50 Å / Num. obs: 10361 / % possible obs: 99.5 % / Redundancy: 5.3 % / Biso Wilson estimate: 65.336 Å2 / CC1/2: 0.986 / Rmerge(I) obs: 0.026 / Rpim(I) all: 0.0289 / Net I/σ(I): 5.24
Reflection shellResolution: 3.5→3.59 Å / Redundancy: 5.5 % / Rmerge(I) obs: 1.261 / Mean I/σ(I) obs: 1.2 / Num. unique obs: 762 / CC1/2: 0.506 / Rrim(I) all: 1.395 / % possible all: 100

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Processing

Software
NameVersionClassification
PHENIX(1.11.1_2575: ???)refinement
XDSMay 1, 2016data reduction
XSCALEMay 1, 2016data scaling
PHASER2.6.0phasing
RefinementMethod to determine structure: MOLECULAR REPLACEMENT
Starting model: PDB entry 4XT1

4xt1


Resolution: 3.5→45.374 Å / SU ML: 0.53 / Cross valid method: FREE R-VALUE / σ(F): 1.34 / Phase error: 28.28
RfactorNum. reflection% reflectionSelection details
Rfree0.2962 1041 10.08 %random selection
Rwork0.252 ---
obs0.2563 10332 99.35 %-
Solvent computationShrinkage radii: 0.9 Å / VDW probe radii: 1.11 Å
Refinement stepCycle: LAST / Resolution: 3.5→45.374 Å
ProteinNucleic acidLigandSolventTotal
Num. atoms4755 0 152 0 4907
Refine LS restraints
Refine-IDTypeDev idealNumber
X-RAY DIFFRACTIONf_bond_d0.0035029
X-RAY DIFFRACTIONf_angle_d0.7016848
X-RAY DIFFRACTIONf_dihedral_angle_d10.1162923
X-RAY DIFFRACTIONf_chiral_restr0.04788
X-RAY DIFFRACTIONf_plane_restr0.004838
LS refinement shell
Resolution (Å)Rfactor RfreeNum. reflection RfreeRfactor RworkNum. reflection RworkRefine-ID% reflection obs (%)
3.5002-3.68470.33011470.321310X-RAY DIFFRACTION100
3.6847-3.91540.37361510.29871298X-RAY DIFFRACTION99
3.9154-4.21750.351410.27861287X-RAY DIFFRACTION100
4.2175-4.64160.26561580.22811314X-RAY DIFFRACTION100
4.6416-5.31240.29351510.22381321X-RAY DIFFRACTION99
5.3124-6.68960.33491440.27571343X-RAY DIFFRACTION99
6.6896-45.37730.23711490.22621418X-RAY DIFFRACTION98

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