Calmodulin Regulation and Identification of Calmodulin Binding Region of Type-3 Ryanodine Receptor Calcium Release Channel

• Published in: Biochemistry (Easton) 2005-11, Vol.44 (45), p.15074-15081
• Main Authors: Yamaguchi, Naohiro, Xu, Le, Pasek, Daniel A, Evans, Kelly E, Chen, S. R. Wayne, Meissner, Gerhard
• Format: Article
• Language: English
• Subjects: Amino Acid Sequence; Binding Sites; Calmodulin - metabolism; Cell Line; Electric Conductivity; Humans; Molecular Sequence Data; Mutation; Oxidation-Reduction; Protein Isoforms - chemistry; Protein Isoforms - metabolism; Ryanodine Receptor Calcium Release Channel - chemistry; Ryanodine Receptor Calcium Release Channel - genetics; Ryanodine Receptor Calcium Release Channel - metabolism; Sequence Alignment
• ISSN: 0006-2960; 1520-4995
• DOI: 10.1021/bi051251t

Ryanodine receptors (RyRs) are a family of intracellular Ca2+ channels that are regulated by calmodulin (CaM). At low Ca2+ concentrations (1 μM), CaM inhibits all three RyR isoforms. Here we report that the regulation of recombinant RyR3 by CaM is sensitive to redox regulation. RyR3 in the presence of reduced glutathione binds CaM with 10−15-fold higher affinity, at low and high Ca2+ concentrations, compared to in the presence of oxidized glutathione. However, compared to RyR1 assayed at low Ca2+ concentrations under both reducing and oxidizing conditions, CaM binds RyR3 with reduced affinity but activates RyR3 to a greater extent. Under reducing conditions, RyR1 and RyR3 activities are inhibited with a similar affinity at [Ca2+] > 1 μM. Mutagenesis studies demonstrate that RyR3 contains a single conserved CaM binding site. Corresponding amino acid substitutions in the CaM binding site differentially affect CaM binding and CaM regulation of RyR3 and those of the two other isoforms. The results support the suggestion that other isoform dependent regions have a major role in the regulation of RyRs by CaM [Yamaguchi et al. (2004) J. Biol. Chem. 279, 36433−36439].