Identification of residues that determine the absence of a Ca(2+)/myristoyl switch in neuronal calcium sensor-1.
The neuronal calcium sensor (NCS) family of Ca(2+)-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 Ca(2+)/myristoyl switch whereby its N-terminal myristoyl group is sequestered in the Ca(2+)-free form and is exposed on Ca(2+) 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 Ca(2+)/myristoyl switch and is membrane-associated even at low Ca(2+) 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 Ca(2+)/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.
O'Callaghan
Dermott W
DW
Physiological Laboratory, University of Liverpool, Crown Street, Liverpool L69 3BX, United Kingdom.
Burgoyne
Robert D
RD
D000595:Amino Acid Sequence; D002118:Calcium / Q000378:metabolism*; D002135:Calcium-Binding Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D006367:HeLa Cells; D051597:Hippocalcin; D006801:Humans; D008969:Molecular Sequence Data; D019814:Myristic Acid / Q000378:metabolism*; D009419:Nerve Tissue Proteins; D051580:Neuronal Calcium-Sensor Proteins; D009474:Neurons / Q000378:metabolism; D009479:Neuropeptides / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D017434:Protein Structure, Tertiary; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism
D002135:Calcium-Binding Proteins; D009419:Nerve Tissue Proteins; D051580:Neuronal Calcium-Sensor Proteins; D009479:Neuropeptides; D011993:Recombinant Fusion Proteins; C101464:frequenin calcium sensor proteins; D019814:Myristic Acid; D051597:Hippocalcin; D002118:Calcium