Calcium-Mediated Reversal of CaM on the Nav 1.2 IQ Motif: Nested Anti-Parallel Sites

Abstract

Calmodulin (CaM) regulates many signal transduction pathways, binding to both enzymes and intracellular segments of several classes of transmembrane receptors and channels. CaM binds its targets with dissociation constants ranging from pM to µM. CaM regulates the inactivation of the neuronal voltage-gated sodium channel 1.2 (NaV1.2). CaM is considered to be a constitutive subunit: both apo and calcium-saturated C-domain of CaM bind tightly to the IQ motif located in the carboxyl-terminal tail of the channel. Apo CaM binds in a semi-open conformation to the IQ motifs of NaV1.2, 1.5, and 1.6. Calcium-saturated CaM binds to the IQ motif, but also binds to a linker between transmembrane domains III and IV. The calcium-dependent competition for CaM between the DIII-DIV linker and the IQ motif is not well understood. Particularly perplexing is the high affinity of CaM for the IQ motif under both apo and calcium-saturating conditions. Does CaM dissociate to bind the III-IV linker? Could a second CaM bind to the channel? How does CaM change position on the channel after binding calcium? To address these questions, we solved the solution (NMR) structure of calcium-saturated CaM C-domain bound to NaV1.2IQ (2M5E). It reveals that calcium binding triggers a reversal relative to that of bound apo CaM C-domain (2KXW). Several NaV1.2IQ residues participate in the interface of both complexes, but their contacts with CaM are distinct because of differences in the topography of the semi-open and open hydrophobic clefts. Comparison of 2M5E with several other complexes of calcium-saturated CaM C-domain bound to target sequences emphasizes why CaM-binding domains are characterized by so many different sequence “motifs”. Favorable energies of association are determined by noncovalent interactions that are made on an atom-by-atom, rather than residue-by-residue, basis. NIH R01 GM57001