Abstract
CaBP4 modulates Ca(2+)-dependent activity of L-type voltage-gated Ca(2+) channels (Cav1.4) in retinal photoreceptor cells. Mg(2+) binds to the first and third EF-hands (EF1 and EF3), and Ca(2+) binds to EF1, EF3, and EF4 of CaBP4. Here we present NMR structures of CaBP4 in both Mg(2+)-bound and Ca(2+)-bound states and model the CaBP4 structural interaction with Cav1.4. CaBP4 contains an unstructured N-terminal region (residues 1-99) and four EF-hands in two separate lobes. The N-lobe consists of EF1 and EF2 in a closed conformation with either Mg(2+) or Ca(2+) bound at EF1. The C-lobe binds Ca(2+) at EF3 and EF4 and exhibits a Ca(2+)-induced closed-to-open transition like that of calmodulin. Exposed residues in Ca(2+)-bound CaBP4 (Phe(137), Glu(168), Leu(207), Phe(214), Met(251), Phe(264), and Leu(268)) make contacts with the IQ motif in Cav1.4, and the Cav1.4 mutant Y1595E strongly impairs binding to CaBP4. We conclude that CaBP4 forms a collapsed structure around the IQ motif in Cav1.4 that we suggest may promote channel activation by disrupting an interaction between IQ and the inhibitor of Ca(2+)-dependent inactivation domain.
Highlights
Cav1.4 is regulated by CaBP4, which is required for continuous release of neurotransmitter in retinal photoreceptor cells
We propose that CaBP4 activates Cav1.4 by binding to the IQ motif, which we suggest may disrupt an interaction between the IQ and inhibitor of Ca2ϩ-dependent inactivation (ICDI) domains
At saturating Ca2ϩ levels, the N-lobe of CaBP4 adopts a closed conformation with Ca2ϩ bound at EF1, in contrast to the Ca2ϩ-bound open conformation seen in CaM [17] and TnC [51]
Summary
Cav1.4 is regulated by CaBP4, which is required for continuous release of neurotransmitter in retinal photoreceptor cells. We conclude that CaBP4 forms a collapsed structure around the IQ motif in Cav1.4 that we suggest may promote channel activation by disrupting an interaction between IQ and the inhibitor of Ca2؉-dependent inactivation domain. Ca2ϩ-dependent regulation of Cav1.4 mediated by CaBP4 is genetically linked to congenital stationary night blindness [1] Particular mutations in both the CACNA1F gene that encodes the ␣1-subunit of an L-type Ca2ϩ channel (Cav1.4␣) [7,8,9,10] and the CABP4 gene [11, 12] were associated with this autosomal recessive retinopathy. In contrast to CaM, CaBP4 contains ϳ100 non-conserved amino acids upstream of the EF-hands in the N-terminal region Another distinguishing property of CaBP4 is that the second EF-hand lacks critical residues required for high affinity Ca2ϩ binding [15, 18]. We propose that CaBP4 activates Cav1.4 by binding to the IQ motif, which we suggest may disrupt an interaction between the IQ and ICDI domains
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