Interactions of N-and C-lobes of Calmodulin with the C-terminal tail of Cav1.2 Ca²⁺ Channel

Abstract

Calmodulin (CaM) is one of the important regulators of the Cav1.2 Ca²⁺ channel. CaM binding to the C-terminal tail of the α1-subunit of the Cav1.2 channel has been shown to be crucial for both Ca²⁺-dependent facilitation (CDF) and Ca²⁺-dependent inactivation (CDI). CaM consists of N-terminal and C-terminal lobes (N-lobe and C-lobe), each containing two Ca²⁺-binding EF-hand regions. However, it is unclear which lobe interacts with the channel during CDF and CDI. In this study, we have examined binding of N-and C-lobes of CaM to the C-terminal tail of Cav1.2 by using the pull-down method. Glutathione-S-transferase (GST)-fusion proteins of N-and C-lobes of CaM and C-terminal peptides containing CB domain (GST-CB) and IQ-motif region (GST-IQ) were prepared, and the GST regions of the fused N-and C-lobes was removed by cleavage. Both N-and C-lobes of CaM bound to GST-IQ in Ca²⁺-dependent manners. We have also examined binding of CaM lobes to GST-CB and other fragments of the C-terminal tail. Possible interactions between N-and C-lobes for the binding to the sites in the channel, and hypothetical conformation model for CDF and CDI will be discussed. [J Physiol Sci. 2008;58 Suppl:S72]