Apo-Calmodulin Binds with its C-terminal Domain to the N-Methyl-D-aspartate Receptor NR1 C0 Region

Calmodulin (CaM) is the major Ca²⁺ sensor in eukaryotic cells. It consists of four EF-hand Ca²⁺ binding motifs, two in its N-terminal domain and two in its C-terminal domain. Through a negative feedback loop, CaM inhibits Ca²⁺ influx through N-methyl-D-aspartate-type glutamate receptors in neurons by binding to the C0 region in the cytosolic tail of the NR1 subunit. Ca²⁺-depleted (apo)CaM is pre-associated with a variety of ion channels for fast and effective regulation of channel activities upon Ca²⁺ influx. Using the NR1 C0 region for fluorescence and circular dichroism spectroscopy studies we found that not only Ca²⁺-saturated CaM but also apoCaM bound to NR1 C0. In vitro interaction assays showed that apoCaM also binds specifically to full-length NR1 solubilized from rat brain and to the complete C terminus of the NR1 splice form that contains the C0 plus C2′ domain. The Ca ²⁺-independent interaction of CaM was also observed with the isolated C- but not N-terminal fragment of calmodulin in the independent spectroscopic assays. Fluorescence polarization studies indicated that apoCaM associated via its C-terminal domain with NR1 C0 in an extended conformation and collapsed to adopt a more compact conformation of faster rotational mobility in its complex with NR1 C0 upon addition of Ca²⁺. Our results indicate that apoCaM is associated with NR1 and that the complex of CaM bound to NR1 C0 undergoes a dramatic conformational change when Ca²⁺ binds to CaM.