Five Members of a Novel Ca2+-binding Protein (CABP) Subfamily with Similarity to Calmodulin (original) (raw)

2000, Journal of Biological Chemistry

Five members of a novel Ca 2؉-binding protein subfamily (CaBP), with 46-58% sequence similarity to calmodulin (CaM), were identified in the vertebrate retina. Important differences between these Ca 2؉-binding proteins and CaM include alterations within their second EF-hand loop that render these motifs inactive in Ca 2؉ coordination and the fact that their central ␣-helixes are extended by one ␣-helical turn. CaBP1 and CaBP2 contain a consensus sequence for N-terminal myristoylation, similar to members of the recoverin subfamily and are fatty acid acylated in vitro. The patterns of expression differ for each of the various members. Expression of CaBP5, for example, is restricted to retinal rod and cone bipolar cells. In contrast, CaBP1 has a more widespread pattern of expression. In the brain, CaBP1 is found in the cerebral cortex and hippocampus, and in the retina this protein is found in cone bipolar and amacrine cells. CaBP1 and CaBP2 are expressed as multiple, alternatively spliced variants, and in heterologous expression systems these forms show different patterns of subcellular localization. In reconstitution assays, CaBPs are able to substitute functionally for CaM. These data suggest that these novel CaBPs are an important component of Ca 2؉-mediated cellular signal transduction in the central nervous system where they may augment or substitute for CaM. Among organisms as diverse as yeast and human, changes in the intracellular Ca 2ϩ ion concentration initiate an array of signaling pathways. Ca 2ϩ ions function as a diffusible signal that exerts its effect directly or through Ca 2ϩ-binding proteins on plasma membrane and intracellular channels, intracellular proteins involved in membrane trafficking, and a broad range of enzymes, including kinases, phosphatases, and adenylyl cyclases. Ca 2ϩ-binding proteins sense changes in [Ca 2ϩ ] through either 130-amino acid (aa) 1 structural elements called C2 domains, 29-aa EF-hand motifs, or through acidic regions of proteins or protein-lipid interfaces. In a growing number of eukaryotic signaling proteins, C2 and EF-hand motifs are present as either a single copy or clustered in multiple copies (1). The largest group of Ca 2ϩ-binding proteins belongs to the calmodulin (CaM) superfamily. They are structurally related and comprise four EF-hand motifs, some of which (one or two) may be nonfunctional in Ca 2ϩ coordination (2). Neuronal Ca 2ϩbinding proteins (NCBP) are a subset of the EF-hand-containing proteins, whose function is largely unknown. The sequence similarity among members of the NCBP family varies from ϳ25% between CaM and visinin to ϳ60% between GCAP1 and GCAP3 (3). NCBPs are acidic and similar in length. CaM and CaM-like proteins are the shortest (149-150 aa; molecular mass, 16,837 Da); other members of this family are ϳ200 aa long (molecular mass, ϳ23,000 Da) (2). NCBPs also display a variety of interesting structural features. Multifunctional CaM contains a pair of N-terminal (EFhand 1 and EF-hand 2) and C-terminal EF-hand (EF-hand 3 and EF-hand 4) motifs (4). The "dumbbell shape" of CaM undergoes a major conformational change upon Ca 2ϩ coordination. These conformational changes result in reorientation of the two N-and C-terminal domains with respect to each other and a rearrangement of ␣-helices in the N-terminal domain that makes the hydrophobic target peptide binding site more accessible (5). The flexible region of the central ␣-helix bends and unwinds upon binding of CaM to many target proteins (6). Myristoylated recoverin, hippocalcin, frequenin, GCAPs, and * This research was supported by National Institutes of Health Grants EY08061 (to K. P.