Relative ligand binding to small or large aggregates measured by scanning correlation spectroscopy

Area of Science:

• Cell Biology
• Biophysics

Background:

• Cell surface receptors mediate cellular activity through signal transduction.
• Ligand-induced receptor aggregation is a key mechanism in receptor activation.
• Receptor systems can exhibit both high and low ligand binding affinities, with models linking clusters to specific affinities.

Purpose of the Study:

• To investigate the relationship between ligand binding and receptor aggregation state.
• To test the hypothesis of preaggregated receptors in 3T3 Swiss mouse fibroblasts.
• To elucidate the binding characteristics of succinyl concanavalin A to cell surface receptors.

Main Methods:

• Utilized scanning fluorescence correlation spectroscopy (sFCS) to analyze receptor dynamics.
• Measured fluorescence fluctuation variance (g[0]) across the cell surface.
• Employed computer simulations to validate experimental findings and control for background fluorescence.

Main Results:

• Fluorescence fluctuation variance (g[0]) increased with varying ligand concentrations (0.33 to 67 mg/L).
• g(0) values plateaued at ligand concentrations above approximately 10 mg/L.
• Observed data are inconsistent with homogeneous receptor distribution or equal binding affinities.

Conclusions:

• The findings support a model of partly aggregated receptors with distinct binding affinities.
• High-affinity binding is associated with small receptor aggregates.
• Low-affinity binding is linked to larger receptor aggregates, explaining the observed ligand concentration-dependent effects.