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Updated: Feb 15, 2026
Studying the Stoichiometry of Epidermal Growth Factor Receptor in Intact Cells using Correlative Microscopy
Published on: September 11, 2015
Investigation of the binding characteristics between ligands and epidermal growth factor receptor by cell membrane
Liu Yang1, Man Zhu1, Yuan Kang1
1School of Pharmacy, Health Science Center, Xi'an Jiaotong University, Xi'an, Shaanxi Province, PR China.
Abstract:
The binding property between a ligand and its receptor is very important for numerous biological processes. In this study, we developed a high epidermal growth factor receptor (EGFR)-expression cell membrane chromatography (CMC) method to investigate the binding characteristics between EGFR and the ligands gefitinib, erlotinib, canertinib, afatinib, and vandetanib. Competitive binding analysis using gefitinib as the marker was used to investigate the interactions that occurred at specific binding sites on EGFR. The ability of displacement was measured from the HEK293-EGFR/CMC column on the binding sites occupied by gefitinib for these ligands, which revealed the following order: gefitinib (KD, 8.49 ± 0.11 × 10-7 M) > erlotinib (KD, 1.07 ± 0.02 × 10-6 M) > canertinib (KD, 1.41 ± 0.07 × 10-6 M) > afatinib (KD, 1.80 ± 0.12 × 10-6 M) > vandetanib (KD, 1.99 ± 0.03 × 10-6 M). This order corresponded with the values estimated by frontal displacement analysis and the scores obtained with molecular docking. Furthermore, thermodynamic analysis indicated that the hydrogen bond or Van der Waals force was the main interaction force in the process of EGFR binding to all 5 ligands. Overall, these results demonstrate that a CMC method could be an effective tool to investigate the binding characteristics between ligands and receptors.
Insights
A new cell membrane chromatography method effectively characterized binding affinities between epidermal growth factor receptor (EGFR) and five cancer drugs. Gefitinib showed the strongest binding, with hydrogen bonds and Van der Waals forces driving interactions.
Area of Science:
- Biochemistry
- Pharmacology
- Analytical Chemistry
Background:
- Ligand-receptor binding is crucial for biological processes.
- Understanding these interactions is key for drug development.
Purpose of the Study:
- To develop and validate a cell membrane chromatography (CMC) method for assessing epidermal growth factor receptor (EGFR) ligand binding.
- To quantify the binding affinities of gefitinib, erlotinib, canertinib, afatinib, and vandetanib to EGFR.
Main Methods:
- High EGFR-expression cell membrane chromatography (CMC) was employed.
- Competitive binding assays using gefitinib as a marker were performed.
- Frontal displacement analysis and molecular docking were utilized for validation.
Main Results:
- The binding affinity order was determined as gefitinib > erlotinib > canertinib > afatinib > vandetanib.
- Dissociation constants (KD) were quantified for each ligand.
- Thermodynamic analysis revealed hydrogen bonds and Van der Waals forces as primary interaction mechanisms.
Conclusions:
- The developed CMC method is effective for investigating ligand-receptor binding characteristics.
- This method provides valuable insights into the binding profiles of EGFR-targeting drugs.
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