Impact of calmodulin missense variants associated with congenital arrhythmia on the thermal stability and the degree of unfolding

Thermal denaturation profiles of proteins that bind several ligands may deviate from the single transition, making their thermodynamic description challenging. We report an empirical method that estimates melting temperatures (Tm) from multi-transition thermal denaturation profiles of 16 variants of calmodulin (CaM) associated with congenital arrhythmia. Differences in Tm estimated by empirical fitting correlate (for apo CaM variants) with those obtained by thermodynamic models. Most CaM variants were more stable than the wild type (WT) in the absence of Ca2+, but less stable in the presence of Ca2+, and displayed either WT-like or higher unfolding percentages in their apo-form, as evaluated by circular dichroism spectroscopy.