Label-Free Assay for Thermodynamic Analysis of Protein−Ligand Interactions: A Multivariate Strategy for Allosteric Ligand Screening

The binding of allosteric ligands to protein can induce changes to the holoprotein conformation, stability, and activity that have an impact on unfolding dynamics. Herein we report a label-free strategy for determining the dissociation constant of protein-ligand interactions over a wide dynamic range (>10(4), Kd from nano- to millimolar) using capillary electrophoresis that overcomes the constraints of an ideal two-state protein unfolding …