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8w2v

Solution Structure of the CD28 hinge used in chimeric antigen receptor (CAR) T-cells

Method: SOLUTION NMR Dmax: 76.1 Å Quality: REASONABLE
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1. Protein Identity and Related Structures Protein Identity & Related Structures

T-cell-specific surface glycoprotein CD28

Homo sapiens

UniProt P10747

State in the Current Structure

Assembly Oligomeric State Construct Mutations and Modifications Ligands, Ions and Associated Components Method and Experimental Conditions Structure Quality
1 Protein monomer Monomer Protein × 1 PDB declaration: monomeric（1） Consistent with protein copy count Chain A； UniProt 114–152 Not recorded No other associated polymer SOLUTION NMR NMR measurement conditions：pH 6.5；298 K；Ionic strength (raw mmCIF value) 70；Pressure 1 NMR measurement conditions：pH 6.5；284 K；Ionic strength (raw mmCIF value) 70；Pressure 1 NMR sample composition：0.5 mM [U-100% 13C; U-100% 15N] CD28 hinge, 20 mM sodium phosphate, 50 mM sodium chloride, 2 mM DTT, 20 uM zinc sulphate, 1 mM pefabloc, 0.1 % sodium azide, 95% H2O/5% D2O | 95% H2O/5% D2O NMR sample composition：0.6 mM [U-100% 13C; U-100% 15N] CD28 hinge, 20 mM sodium phosphate, 50 mM sodium chloride, 2 mM DTT, 20 uM zinc sulphate, 1 mM pefabloc, 0.1 % sodium azide, 100% D2O | 100% D2O NMR sample composition：0.45 mM [U-100% 13C; U-100% 15N] CD28 hinge, 20 mM sodium phosphate, 50 mM sodium chloride, 2 mM DTT, 20 uM zinc sulphate, 1 mM pefabloc, 0.1 % sodium azide, 95% H2O/5% D2O | 95% H2O/5% D2O NMR sample composition：0.4 mM [U-100% 15N] CD28 hinge, 20 mM sodium phosphate, 50 mM sodium chloride, 2 mM DTT, 20 uM zinc sulphate, 1 mM pefabloc, 0.1 % sodium azide, 95% H2O/5% D2O | 95% H2O/5% D2O NMR sample composition：0.28 mM [U-100% 15N] CD28 hinge, 20 mM sodium phosphate, 50 mM sodium chloride, 2 mM DTT, 20 uM zinc sulphate, 1 mM pefabloc, 0.1 % sodium azide, 95% H2O/5% D2O | 95% H2O/5% D2O Resolution not provided

Other States of the Same Protein in the Database

Each row is a biological assembly of the same UniProt protein in another PDB entry. The “Difference from current entry” column identifies evidence-level differences; no tag means the currently parsed fields agree.

Other PDB Difference from Current Entry 8W2V Assembly / Oligomeric State Construct Mutations and Modifications Ligands, Ions and Non-polymers Method and Experimental Conditions Structure Quality
1YJD Crystal structure of human CD28 in complex with the Fab fragment of a mitogenic antibody (5.11A1) Deposited 2005-01-14 Different construct Different oligomeric state Different ligand/ion Different experimental method Different experimental conditions Different structure-quality metrics Assembly 1 Protein heterocomplex Heteromer；Protein × 6 PDB declaration: hexameric
Chain C 17–152（136 aa） Fragment：Extracellular region
Not recorded NAG 2-acetamido-2-deoxy-beta-D-glucopyranose × 6 X-RAY DIFFRACTION
X-ray crystallization conditions VAPOR DIFFUSION, SITTING DROP；pH 5.9；294 K；PEG 3350, magnesium formate, pH 5.9, VAPOR DIFFUSION, SITTING DROP, temperature 294K
Resolution 2.70 Å R-free 0.282
3WA4 Grb2 SH2 domain/CD28-derived peptide complex Deposited 2013-04-23 Different construct Different mutation/modification Different experimental method Different experimental conditions Different structure-quality metrics Assembly 2 Protein monomer Monomer；Protein × 1 PDB declaration: monomeric
Chain B 189–196（8 aa） Fragment：fragment peptide, UNP residues 190-195
Non-standard monomer：Yes (specific site not provided by mmCIF) No recorded non-water small molecule X-RAY DIFFRACTION
X-ray crystallization conditions VAPOR DIFFUSION, HANGING DROP；pH 7.5；293 K；0.1M HEPES, 1.25M sodium acetate, 0.1M cadmium sulfate, pH 7.5, VAPOR DIFFUSION, HANGING DROP, temperature 293K
Resolution 1.35 Å R-free 0.209
3WA4 Grb2 SH2 domain/CD28-derived peptide complex Deposited 2013-04-23 Different construct Different mutation/modification Different oligomeric state Different ligand/ion Different experimental method Different experimental conditions Different structure-quality metrics Assembly 3 Protein heterocomplex Heteromer；Protein × 2 PDB declaration: dimeric
Chain B 189–196（8 aa） Fragment：fragment peptide, UNP residues 190-195
Non-standard monomer：Yes (specific site not provided by mmCIF) CD CADMIUM ION × 1 ACY ACETIC ACID × 1 X-RAY DIFFRACTION
X-ray crystallization conditions VAPOR DIFFUSION, HANGING DROP；pH 7.5；293 K；0.1M HEPES, 1.25M sodium acetate, 0.1M cadmium sulfate, pH 7.5, VAPOR DIFFUSION, HANGING DROP, temperature 293K
Resolution 1.35 Å R-free 0.209
5AUL PI3K p85 C-terminal SH2 domain/CD28-derived peptide complex Deposited 2015-04-28 Different construct Different mutation/modification Different oligomeric state Different ligand/ion Different experimental method Different experimental conditions Different structure-quality metrics Assembly 1 Protein heterocomplex Heteromer；Protein × 2 PDB declaration: dimeric
Chain B 189–196（8 aa） Fragment：UNP residues 189-196
Non-standard monomer：Yes (specific site not provided by mmCIF) GOL GLYCEROL × 1 X-RAY DIFFRACTION
X-ray crystallization conditions VAPOR DIFFUSION, HANGING DROP；293 K；100 mM Sodium Acetate trihydrate, 200 mM Ammonium Acetate, 30% PEG4000
Resolution 1.10 Å R-free 0.160
5GJH Gads SH2 domain/CD28-derived peptide complex Deposited 2016-06-30 Different construct Different mutation/modification Different oligomeric state Different experimental method Different experimental conditions Different structure-quality metrics Assembly 1 Protein heterocomplex Heteromer；Protein × 2 PDB declaration: dimeric
Chain B 189–196（8 aa） Fragment：UNP residues 189-196
Non-standard monomer：Yes (specific site not provided by mmCIF) No recorded non-water small molecule X-RAY DIFFRACTION
X-ray crystallization conditions VAPOR DIFFUSION, HANGING DROP；pH 7.5；293 K；100 mM HEPES-NaOH, 10% PEG 6000, 20% MPD
Resolution 1.20 Å R-free 0.170
5GJH Gads SH2 domain/CD28-derived peptide complex Deposited 2016-06-30 Different construct Different mutation/modification Different oligomeric state Different experimental method Different experimental conditions Different structure-quality metrics Assembly 2 Protein heterocomplex Heteromer；Protein × 2 PDB declaration: dimeric
Chain D 189–196（8 aa） Fragment：UNP residues 189-196
Non-standard monomer：Yes (specific site not provided by mmCIF) No recorded non-water small molecule X-RAY DIFFRACTION
X-ray crystallization conditions VAPOR DIFFUSION, HANGING DROP；pH 7.5；293 K；100 mM HEPES-NaOH, 10% PEG 6000, 20% MPD
Resolution 1.20 Å R-free 0.170
5GJI PI3K p85 N-terminal SH2 domain/CD28-derived peptide complex Deposited 2016-06-30 Different construct Different mutation/modification Different oligomeric state Different ligand/ion Different experimental method Different experimental conditions Different structure-quality metrics Assembly 1 Protein heterocomplex Heteromer；Protein × 2 PDB declaration: dimeric
Chain B 189–196（8 aa） Fragment：UNP residues 189-196
Non-standard monomer：Yes (specific site not provided by mmCIF) SO4 SULFATE ION × 3 GOL GLYCEROL × 1 X-RAY DIFFRACTION
X-ray crystallization conditions VAPOR DIFFUSION, HANGING DROP；pH 4.6；293 K；100mM Sodium acetate trihydrate, 30% PEG MME 2000, 200mM Ammonium sulfate
Resolution 0.90 Å R-free 0.145
6O8D Anti-CD28xCD3 CODV Fab bound to CD28 Deposited 2019-03-09 Different construct Different oligomeric state Different ligand/ion Different experimental method Different experimental conditions Different structure-quality metrics Assembly 1 Other combination Heteromer；Protein × 6 PDB declaration: hexameric
Chain C 33–150（118 aa）
Not recorded NAG 2-acetamido-2-deoxy-beta-D-glucopyranose × 2 X-RAY DIFFRACTION
X-ray crystallization conditions VAPOR DIFFUSION, SITTING DROP；277 K；26% PEG3350, 0.75% myo-inositol, 0.263M potassium citrate tribasic monohydrate
Resolution 3.55 Å R-free 0.356
7PPN SHP2 catalytic domain in complex with CD28 (183-198) phosphopeptide (pTyr-191, p-Thr-195) Deposited 2021-09-14 Different construct Different mutation/modification Different oligomeric state Different ligand/ion Different experimental method Different experimental conditions Different structure-quality metrics Assembly 1 Protein heterocomplex Heteromer；Protein × 2 PDB declaration: dimeric
Chain B 183–198（16 aa）
Non-standard monomer：Yes (specific site not provided by mmCIF) GOL GLYCEROL × 1 X-RAY DIFFRACTION
X-ray crystallization conditions VAPOR DIFFUSION, HANGING DROP；pH 5.5；277 K；10% PEG 20000, 100 mM Citrate buffer pH 5.5, 50 mM EDTA
Resolution 1.90 Å R-free 0.207
7VU5 Structure of the transmembrane domain of the CD28 dimer Deposited 2021-11-01 Different construct Different mutation/modification Different oligomeric state Different experimental conditions Assembly 1 Protein homooligomer Homooligomer；Protein × 2 PDB declaration: dimeric
Chain A 148–188（41 aa）
Chain B 148–188（41 aa）
Not recorded No recorded non-water small molecule SOLUTION NMR
NMR measurement conditions pH 6；310 K；Ionic strength (raw mmCIF value) 25；Pressure 1
NMR sample composition 0.6 mM U-13C; U-15N; 75%-2H CD28_TM, 25 mM MES, 30 mM DMPC, 60 mM DHPC, 90% H2O/10% D2O | 90% H2O/10% D2O
NMR sample composition 0.6 mM [U-13C; U-15N] CD28_TM, 25 mM MES, 30 mM acyl chain U-2H DMPC, 60 mM acyl chain U-2H DHPC, 90% H2O/10% D2O | 90% H2O/10% D2O
NMR sample composition 0.6 mM U-15N; U-2H CD28_TM, 0.6 mM [U-13C] CD28_TM, 25 mM MES, 30 mM acyl chain U-2H DMPC, 60 mM acyl chain U-2H DHPC, 90% H2O/10% D2O | 90% H2O/10% D2O
NMR sample composition 0.2 mM U-15N; 15%-13C CD28_TM, 25 mM MES, 30 mM acyl chain U-2H DMPC, 60 mM acyl chain U-2H DHPC, 90% H2O/10% D2O | 90% H2O/10% D2O
Resolution not provided
8S6Z CD28 in complex with the antibody Fab fragment AI3 Deposited 2024-02-28 Different construct Different oligomeric state Different ligand/ion Different experimental method Different experimental conditions Different structure-quality metrics Assembly 1 Other combination Heteromer；Protein × 3 PDB declaration: trimeric
Chain C 19–152（134 aa）
Not recorded IMD IMIDAZOLE × 4 GOL GLYCEROL × 3 ZN ZINC ION × 13 NAG 2-acetamido-2-deoxy-beta-D-glucopyranose × 4 X-RAY DIFFRACTION
X-ray crystallization conditions VAPOR DIFFUSION, SITTING DROP；pH 7.4；293 K；Crystallization trials were set up using the CD28-AI3 Fab at 15.8 mg/mL in phosphate buffered saline (PBS) pH 7.4. Drops were set up using 100 nl protein and 100 nL reservoir solution containing 200 mM zinc acetate, 0.1 M imidazole pH 8.0, 20% w/v PEG 3000.
Resolution 3.05 Å R-free 0.258
8S6Z CD28 in complex with the antibody Fab fragment AI3 Deposited 2024-02-28 Different construct Different oligomeric state Different ligand/ion Different experimental method Different experimental conditions Different structure-quality metrics Assembly 2 Protein heterocomplex Heteromer；Protein × 3 PDB declaration: trimeric
Chain F 19–152（134 aa）
Not recorded IMD IMIDAZOLE × 3 GOL GLYCEROL × 3 ZN ZINC ION × 13 NAG 2-acetamido-2-deoxy-beta-D-glucopyranose × 5 X-RAY DIFFRACTION
X-ray crystallization conditions VAPOR DIFFUSION, SITTING DROP；pH 7.4；293 K；Crystallization trials were set up using the CD28-AI3 Fab at 15.8 mg/mL in phosphate buffered saline (PBS) pH 7.4. Drops were set up using 100 nl protein and 100 nL reservoir solution containing 200 mM zinc acetate, 0.1 M imidazole pH 8.0, 20% w/v PEG 3000.
Resolution 3.05 Å R-free 0.258

9 other PDB entries and 12 assemblies. Open the comparison page and filter oligomeric states

View Construct and Data Evidence
UniProt name CD28_HUMAN
Isoform —
PDB entities 1
Chains and sequence ranges Author chain A； PDBConstruct 1–39； UniProt 114–152

The page prioritizes protein identity, the current assembly, associated components, oligomeric state and cross-PDB links. Chain mapping and sequence ranges are retained as data evidence. Internal IDs, import timestamps and assembly operation expressions are maintenance fields and are not shown here.

SAXS scattering curve SAXS Profile

SAXS profile for 8w2v

P(r) Distance Distribution P(r) Distribution

P(r) distribution for 8w2v
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2. Structure Basics 2. Structure Basics

Entry ID entry_id8w2v
Deposition date deposition_date2024-02-21
最后修订 last_revision2024-09-11
Structure title titleSolution Structure of the CD28 hinge used in chimeric antigen receptor (CAR) T-cells
Keywords keywordsCAR-T, CD28, immunotherapy, intrinsic disorder, IMMUNE SYSTEM; IMMUNE SYSTEM
Experimental Method methodSOLUTION NMR
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3. SAXS Parameters (CRYSOL theoretical calculation) 3. SAXS Parameters (CRYSOL)

Radius of gyration Rg (Guinier) rg_guinier19.59 Å
Radius of gyration Rg (electron density) rg_electron19.66 Å
Forward intensity I(0) i0155021000.00
Molecular weight molecular_weight107580.0 kDa
Excluded volume excluded_volume137370 ų
Envelope volume envelope_volume91779 ų
Hydration-shell volume shell_volume30685 ų
Envelope diameter envelope_diameter87.7 Å
Shell Rg shell_rg32.21 Å
Envelope Rg envelope_rg25.00 Å
Shape Rg shape_rg19.62 Å
Total Rg total_rg20.82 Å
Total atoms total_atoms15325
Residues n_residues975
Spherical-harmonic order n_harmonics20
q range q_range— – 0.5000 Å−1
Data points n_points101
Shell type shell_typedirectional
Solvent electron density solvent_density0.3340 e/ų
Shell contrast contrast_shell0.0300 e/ų
CRYSOL version crysol_version4.1.3
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4. P(r) Distance Distribution (GNOM inversion) 4. P(r) Analysis (GNOM)

Maximum dimension Dmax dmax76.1 Å
Rg (real space) rg_real19.77 Å
Rg uncertainty (real space) rg_real_error1.02 Å
I(0) (real space) i0_real1.5500e+08
I(0) uncertainty (real space) i0_real_error2.3880e+06
Rg (reciprocal space) rg_reciprocal19.75 Å
I(0) (reciprocal space) i0_reciprocal155000000.0000
Solution quality estimate total_estimate0.5852
Solution quality rating solution_quality REASONABLE a REASONABLE solution
P(r) peaks n_peaks3
Primary peak position r_peak_primary22.5 Å
Skewness Skewness skewness0.452
Kurtosis Kurtosis kurtosis-0.056
Angular range angular_range— – 0.4050 Å−1
Current regularization parameter α current_alpha0.0000
Highest regularization parameter α highest_alpha121000.0000
Real-space data points n_real_points72
GNOM version gnom_version4.1.3
Quality Criteria quality_criteria AN1: 0.000; Oscil: 0.588; Stabil: 1.000; Sysdev: 0.424; Positv: 1.000; Valcen: 0.567; Smooth: 0.000
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5. Crystallography and Experiment 5. Crystallography & Experiment

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6. Entities and Polymers Entities & Polymers (1)

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8. Citations (1)

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9. Files and Curves (10)