1. Investigation of Dye-Protein Interactions and Optimization of Fluorescence-Based Assays for Target Binding of Calmodulin
Carey K. Johnson, Department of Chemistry, University of Kansas, Lawrence, KS 66045
Fluorescence polarization assays detect the change in rotational mobility of a tracer (here calmodulin) upon binding to a target. +
CaM-fl PMCA CaM-fl:PMCA
Low Anisotropy
High Anisotropy
We identified a dye label, Atto 465, that sticks to calmodulin, to increase the sensitivity of the fluorophore to target binding. Fluorescence polarization measurements with CaM-Atto 465 yields the binding affinity (measured directly for the first time). We are currently developing a competition assay for binding of calmodulin to peptides.