Urea-induced denaturation of human calcium/calmodulin-dependent protein kinase IV: a combined spectroscopic and MD simulation studies

CaMKIV plays important role in varieties of biological processes such as gene expression regulation, memory consolidation, bone growth, T-cell maturation, sperm motility, regulation of microtubule dynamics, cell-cycle progression, and apoptosis. To measure stability parameters, urea-induced denaturation of CaMKIV was carried out at pH 7.4 and 25°C, using three different probes, namely far-UV CD, near-UV absorption, and tryptophan fluorescence.

A coincidence of normalized denaturation curves of these optical properties suggests that urea-induced denaturation is a two-state process. Analysis of these denaturation curves gave values of 4.20 ± 0.12 kcal mol −1 , 2.95 ± 0.15 M, and 1.42 ± 0.06 kcal mol −1 M −1 for (Gibbs free energy change (Δ G D ) in the absence of urea), C m (molar urea concentration ([urea]) at the midpoint of the denaturation curve), and m (=∂Δ G D /∂[urea]), respectively.