Charged Side Chain Mutations of CaMKII Peptide Alter Binding Affinity for CaM through Creation of Non-local Alternate Binding Contacts

Abstract

Calmodulin (CaM) trapping is a phenomenon where the active and inactive states of calmodulin dependent protein kinase II (CaMKII) produce drastically different affinities for Ca2+ saturated CaM. The states of CaMKII are paramount in the scheme of calcium ion signaling, which is an essential biological function whose underlying mechanism is largely unknown. Experimentally, a set of peptides modeled after CaMKII's binding domain (293-312) were created through systematic mutations of charged residues to mimic the two distinct affinity states for CaM and probe the mechanisms responsible for the observed change in kinetics. Although a model was successfully created, the observed interactions could not be explained through current protein interaction models or electrostatic steering effects. We investigate the dynamics of this experiment through the use of all atom simulations, choosing three of the mutant peptides with a length of 20 amino acids. We refer to these peptides by the 296-298 residues, specifically RRK (wildtype), RAK (1-residue mutation) and AAA (3-residue mutation), and validate our simulation findings through comparison with circular dichroism (CD) data. We demonstrate that the large side chains present in the first 6 residues of each peptide interact with each other, and that the mutation of charged residues produce global changes in sidechain conformations and disordered regions. The specific ensemble of secondary structure/rotamer conformations present in RRK, that are not observed in AAA, are essential for high affinity binding between CaMKII and CaM.