Multiple Actions of Imperatoxin A on Ryanodine Receptors - Interactions with the II-III Loop A Fragment*




Dulhunty, Angela
Curtis, Suzanne
Watson, Sarah
Cengia, Louise
Casarotto, Marco

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American Society for Biochemistry and Molecular Biology Inc


Imperatoxin A is a high affinity activator of ryanodine receptors. The toxin contains a positively charged surface structure similar to that of the A fragment of skeletal dihydropyridine receptors (peptide A), suggesting that the toxin and peptide could bind to a common site on the ryanodine receptor. However, the question of a common binding site has not been resolved, and the concentration dependence of the actions of the toxin has not been fully explored. We characterize two novel high affinity actions of the toxin on the transient gating of cardiac and skeletal channels, in addition to the well documented lower affinity induction of prolonged substates. Transient activity was (a) enhanced with 0.2-10 nM toxin and (b) depressed by >50 nM toxin. The toxin at ≥nM enhanced Ca2+ release from SR in a manner consistent with two independent activation processes. The effects of the toxin on transient activity, as well as the toxin-induced substate, were independent of cytoplasmic Ca2+ or Mg2+ concentrations or the presence of adenine nucleotide and were seen in diisothiocyanostilbene-2′,2′ -disulfonic acid-modified channels. Peptide A activated skeletal and cardiac channels with 100 nM cytoplasmic Ca2+ and competed with Imperatoxin A in the high affinity enhancement of transient channel activity and Ca2+ release from SR. In contrast to transient activity, prolonged substate openings induced by the toxin were not altered in the presence of peptide A. The results suggest that Imperatoxin A has three independent actions on ryanodine receptor channels and competes with peptide A for at least one action.



Keywords: Amino acids; Chemical bonds; Concentration (process); Musculoskeletal system; Proteins; Structure (composition); Cardiac channels; Toxins; Catalyst poisoning; 4,4' diisothiocyanatostilbene 2,2' disulfonic acid; adenine nucleotide; calcium ion; imperatoxin



Journal of Biological Chemistry


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