Effects of peptide C corresponding to the Glu724-Pro760 region of the II-III loop of the DHP (dihydropyridine) receptor α1 subunit on the domain-switch-mediated activation of RyR1 (ryanodine receptor 1) Ca2+ channels

The Leu(720)-Leu(764) region of the II-III loop of the dihydropyridine receptor is believed to be important for both orthograde and retrograde communications with the RyR (ryanodine receptor), but its actual role has not yet been resolved. Our recent Studies suggest that voltage-dependent activation of the RyR channel is mediated by a pair of interacting N-terminal and central domains, designated as the 'domain switch'. To investigate the effect of peptide C (a peptide corresponding to residues Glu(724)-Pro(760)) on domain switch-mediated activation of the RyR, we measured C 121 release induced by DIP (domain peptide) 1 or DP4 (which activates the RyR by mediation of the domain switch) and followed the Ca2+ release time Course using a luminal Ca2+-probe (chlortetracycline) under Ca2+-clamped conditions. Peptide C produced a significant potentiation of the domain-switch-mediated Ca2+ release, provided that the Ca2+ concentration Was sufficiently low (c.g.domain peptide. However, at micromolar Ca2+ concentrations, peptide C inhibits activation. Covalent cross-linking of fluorescently labelled peptide C to the RyR and screening of the fluorescently labelled tryptic fragments permitted LIS to localize the peptide-C-binding site to residues 450-1400, which may represent the primary region involved in physical Coupling. Based oil the above findings, we propose thin the physiological role of residues Glu(724)-Prp(760) is to facilitate depolarization-induced and domain-switch-mediated RyR activation at sub- or near-threshold concentrations of cytoplasmic Ca2+ and to Suppress activation upon all increase of cytoplasmic Ca2+.