Identification of residues that determine the absence of a Ca2+/myristoyl switch in neuronal calcium sensor-1

The neuronal calcium sensor (NCS) family of Ca2+-binding proteins regulates a number of different processes in neurons and photoreceptor cells. The first of these proteins to be characterized, recoverin, was shown to exhibit a Ca2+/myristoyl switch whereby its N-terminal myristoyl group is sequestered in the Ca2+-free form and is exposed on Ca2+ binding to allow the protein to become membrane-associated. It has subsequently been shown that certain other family members also exhibit this mechanism in living cells. In contrast, NCS-1 does not show the Ca2+/myristoyl switch and is membrane-associated even at low Ca2+ concentrations. We have used sequence comparison combined with information from structural analyses to attempt to identify candidate residues within the NCS proteins that determine whether or not the Ca2+/myristoyl switch operates in cells and have tested their functional significance by mutagenesis. The results show that NCS-1 possesses residues within its N terminus that lock the myristoyl group in an exposed conformation. In addition, other structural aspects within the C-terminal domains are required to allow the switch to operate. We have determined a key role for residues within the motif EELTRK in NCS-1 in keeping the myristoyl group exposed and allowing the protein to be constitutively membrane-associated.