The Effect of Calcium Ions and Peptide Ligands on the Relative Stabilities of the Calmodulin Dumbbell and Compact Structures

A combination of ion mobility and mass spectrometry methods was used to characterize the molecular shape of the protein calmodulin (CaM) and its complexes with calcium and a number of peptide ligands. CaM, a calcium-binding protein composed of 148 amino acid residues, was found by X-ray crystallography to occur both in a globular shape and in the shape of an extended dumbbell. Here, it was found, as solutions of Cam and Cam complexes were sprayed into the solvent-free environment of the mass spectrometer, that major structural feature,., of the molecule and the stoichiometry of the units constituting a complex in Solution were preserved in the desolvation process. Two types of Cam structures were observed in Our experiments: a compact and all extended form of Cam with measured cross sections in near-perfect agreement with those calculated for the known globular and extended dumbbell X-ray geometries. Calcium-free Solutions yielded predominantly all extended Cam conformation. Ca-n(2+)-CaM complexes were observed in Calcium-containing solutions, n = 0-4, with the population of the compact conformation increasing relative to the elongated conformation as n increases. For n = 4, a predominantly compact globular conformation was observed. Solutions containing the peptide CaMKII290-309, the CaM target domain of the Ca2+/Calmodulin-dependent protein kinase II (CaMKII) enzyme, yielded predominantly globular Ca-4(2+)-CaM-CaMKII290-309 complexes. Similar results were obtained with the 26-residue peptide melittin. For the 14-residue C-terminal melittin fragment, oil the other hand, formation of both a 1: 1 and a 1:2 CaM-peptide complex was detected. Oil the basis of the entirety of our results, we Conclude that the collapse of extended (dumbbell-like) CaM structures into more compact globular Structures Occurs upon specific binding Of four calcium ions. Furthermore, this calcium-induced structural Collapse of Cam appears to be a prerequisite for formation of a particularly stable CaM-peptide complex involving peptides long enough to be engaged in interactions with both lobes of CaM.