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Published on: August 2, 2018
Heterogeneity in EGF-binding affinities arises from negative cooperativity in an aggregating system
Jennifer L Macdonald1, Linda J Pike
1Department of Biochemistry and Molecular Biophysics, Washington University School of Medicine, 660 South Euclid, Box 8231, St. Louis, MO 63110, USA.
Summary
Epidermal Growth Factor (EGF) receptor binding exhibits negative cooperativity in aggregating systems, explaining multiple binding sites. Receptor expression levels influence signaling outcomes, impacting cellular responses.
Area of Science:
- Biochemistry
- Cell Biology
- Molecular Signaling
Background:
- Scatchard analysis of Epidermal Growth Factor (EGF) binding to its receptor typically shows concave up plots, suggesting two classes of binding sites.
- The molecular basis for these distinct binding sites arising from a single EGF receptor protein has remained unexplained.
Purpose of the Study:
- To develop a new analytical approach to explain the presence of two EGF receptor binding sites.
- To investigate the role of receptor aggregation and negative cooperativity in EGF binding kinetics.
- To analyze the contribution of kinase domain dimers to receptor association and cooperativity.
Main Methods:
- Simultaneous fitting of binding isotherms from cells with varying EGF receptor expression levels.
- Application of a novel model incorporating negative cooperativity within an aggregating system.
- Analysis of EGF receptor mutants, including L680N and Y246D, to probe dimerization and cooperativity mechanisms.
Main Results:
- Binding data is fully explained by a model of negative cooperativity in an aggregating system.
- An experimentally determined monomer-dimer equilibrium constant suggests approximately 50,000 receptors per cell for wild-type EGF receptors.
- Kinase domain dimers contribute significantly to ligand-independent monomer association but are not essential for negative cooperativity.
Conclusions:
- Negative cooperativity in an aggregating system provides a unified explanation for EGF receptor binding heterogeneity.
- Physiological variations in EGF receptor expression levels can modulate cellular responses to signaling stimuli.
- The developed model offers a framework for understanding EGF receptor family dimerization and lateral signaling.
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