The Allostery and Specificity of EF Hand Calcium-Binding Proteins

Abstract

EF-hand Ca2+-binding proteins (CBPs), such as calmodulin (CaM) or those belonging to the S100 protein family (S100s) undergo conformational changes upon increasing intracellular Ca2+, facilitating interactions with protein targets and inducing important biological responses. In the absence of target, the Ca2+-binding affinity of CaM and most S100 proteins is weak (CaKD >1 μM). However, when bound to effectors, allosteric mechanisms increase the Ca2+ affinity of these CBPs (CaKD <1 μM) to allow for proper Ca2+ homeostasis and maintenance of Ca2+-signaling. The Ca2+-tightening of these CBPs is described here by the “binding and functional folding” framework for detailing this physiologically relevant phenomenon. This research seeks to elucidate both the mechanisms of allostery and the basis of ligand- and target- specificity for S100 proteins and CaM. First, molecular fragments were used to differentiate between two highly similar S100 proteins, S100B and S100A1, with the goal of designing S100-specific inhibitors to block these elevated S100 proteins in various disease states. This provided insight into the specificity of S100A1 versus S100B for small molecules and will enable improved S100 protein-based drug design efforts. S100A1 and S100B binding to ions was also compared and differences between ion binding sites within the two highly similar proteins were determined. Another study revealed an allosteric mechanism in which a peptide termed BP2, derived from the STRA6 vitamin A transporter, increased the Ca2+-binding affinity of CaM upon binding. CaM-STRA6 complex formation was observed at physiologically relevant free Ca2+ concentrations (<1 μM), suggesting that retinol transport by full-length CaCaM-STRA6 may be regulated by Ca2+-signaling. The effect of CaM on multiple full-length target proteins was then discussed to further describe CaM allostery. Together, these studies lead to an improved understanding of Ca2+ signaling, CBP allostery and CBP-target- and ligand- interaction specificity.