Background and purpose: Members of the calcin family, presently including imperatoxin A, maurocalcin, opicalcins and hemicalcin, are basic, 33-mer peptide activators of ryanodine receptors (RyRs), the calcium channels of the sarcoplasmic reticulum (SR) that provide the majority of calcium for muscle contraction. Here we describe hadrucalcin, a novel member of this family.
Experimental approach: Hadrucalcin was isolated from the venom of Hadrurus gertschi. Amino acid sequence and mass were determined by Edman degradation and mass spectrometry respectively. A cDNA library was constructed to generate clones for DNA sequence determination. Biological activity of native toxin was confirmed with [(3)H]ryanodine binding, by using SR vesicles from cardiac and skeletal muscle, and with single skeletal (RyR1) and cardiac (RyR2) channels reconstituted in lipid bilayers. Hadrucalcin was applied to intact ventricular myocytes to investigate effects on calcium transients. The secondary structure of hadrucalcin was computer-modelled by using atomic coordinates from maurocalcin, a structurally similar peptide.
Key results: Hadrucalcin is distinguished from previously described congeners by two additional amino acids in its primary sequence and the lack of prominent amphipathicity. Hadrucalcin activated RyRs with high affinity (EC(50)= 37 nmol.L(-1)), induced a long-lasting subconductance state on RyR1 and RyR2, and rapidly (lag time approximately 2 s) penetrated ventricular cardiomyocytes, eliciting discharge of internal calcium stores and spontaneous contractions.
Conclusions and implications: Hadrucalcin is a cell-permeant, powerful activator of RyRs, which has translational potential for targeted delivery of drugs to RyR as novel therapeutic intervention in arrhythmogenic disease.