THE STRUCTURE OF A CALMODULIN MUTANT WITH A DELETION IN THE CENTRAL HELIX - IMPLICATIONS FOR MOLECULAR RECOGNITION AND PROTEIN-BINDING

Background: Calmodulin (CaM) is the major calcium-dependent regulator of a large variety of important intracellular processes in eukaryotes. The structure of CaM consists of two globular calcium-binding domains joined by a central 28-residue a helix. This linker helix has been hy pothesized to act as a flexible tether and is crucial for the binding and activation of numerous target proteins. Although the way in which alterations of the central helix modulate the molecular recognition me chanism is not known exactly, the relative orientation of the globular domains seems to be of great importance. The structural analysis of c entral helix mutants may contribute to a better understanding of how c hanges in the conformation of CaM effect its function. Results: We hav e determined the crystal structure of a calcium-saturated mutant of ch icken CaM (mut-2) that lacks two residues in the central helix, Thr79 and Asp80, at 1.8 Angstrom resolution. The mutated shorter central hel ix is straight, relative to that of the wild-type structure. The loss of a partial turn of the central alpha helix causes the C-terminal dom ain to rotate 220 degrees around the helix axis, with respect to the N -terminal domain. This rotation places the two domains on the same sid e of the central helix, in a cis orientation, rather than in the trans orientation found in wild-type structures. Conclusions: The deletion of two residues in the central helix of CaM does not distort or cause a bending of the linker alpha helix. The main consequence of the mutat ion is a change in the relative orientation of the two globular calciu m-binding domains, causing the hydrophobic patches in these domains to be closer and much less accessible to interact with the target enzyme s. This may explain why this mutant of CaM shows a marked decrease in its ability to activate some enzymes while the mutation has little or no effect on its ability to activate others.