A structural requirement for activation of skeletal ryanodine receptors by peptides of the dihydropyridine receptor II-III loop

The solution structures of three related peptides (A1, A2, and A9) corresponding to the Thr⁶⁷¹-Leu⁶⁹⁰ region of the skeletal muscle dihydropyridine receptor II-III loop have been investigated using nuclear magnetic resonance spectroscopy. Peptide A1, the native sequence, is less effective in activating ryanodine receptor calcium release channels than A2 (Ser⁶⁸⁷ to Ala substitution). Peptide A9, Arg⁶⁸¹-Ser⁶⁸⁷, does not activate ryanodine receptors. A1 and A2 are helical from their N terminus to Lys⁶⁸⁵ but are generally unstructured from Lys⁶⁸⁵ to the C terminus. The basic residues Arg⁶⁸¹-Lys⁶⁸⁵, essential for A1 activation of ryanodine receptors, are located at the C-terminal end of the α-helix. Peptide A9 was found to be unstructured. Differences between A1 and A2 were observed in the C-terminal end of the helix (residues 681-685), which was less ordered in A1, and in the C-terminal region of the peptide, which exhibited greater flexibility in A1. Predicted low energy models suggest that an electrostatic interaction between the hydroxyl oxygen of Ser⁶⁸⁷ and the gnanidino moiety of Arg⁶⁸³ is lost with the Ser⁶⁸⁷Ala substitution. The results show that the more structured peptides are more effective in activating ryanodine receptors.