Expression and Purification of Calmodulin and its Interaction with the SK peptide
Calmodulin (CaM) is a highly conserved protein that has been observed to regulate ion channels. Calcium (Ca2+) binds to Calmodulin in order to activate Ca2+ activated Potassium channels (SK Channels). Calmodulin is known to responsible for gating the SK channel; however, the mechanism in which it does this is unclear. In order to study this interaction, wild-type CaM, single lobe or half CaM, double lobe CaM, and a SK channel peptide (SKp) were all purified. Purification of proteins was done through high performance liquid chromatography (HPLC) and verified with SDS gels. The CaM-SKp complex was analyzed through CG-MALS (composition gradient multi-angle light scattering). CG-MALS can be used to determine the molar mass of complexes formed. Stoichiometry can be determined if molar mass of the proteins complexing are known. Results from CG-MALS show the formation of stoichiometries that have not been observed before. The introduction of these new stoichiometries adds complexity to the SKp-CaM interaction and require more testing in order to fully elucidate the mechanism of SK channel gating.