title: Maurocalcine and peptide A stabilize distinct subconductance states of ryanodine receptor type 1, revealing a proportional gating mechanism.
abstract: Maurocalcine (MCa) isolated from Scorpio maurus palmatus venom shares 82% sequence identity with imperatoxin A. Both scorpion toxins are putative mimics of the II-III loop peptide (termed peptide A (pA)) of alpha(1s)-dihydropyridine receptor and are thought to act at a common site on ryanodine receptor type 1 (RyR1) important for skeletal muscle EC coupling. The relationship between the actions of synthetic MCa (sMCa) and pA on RyR1 were examined. sMCa released Ca(2+) from SR vesicles (EC(50) = 17.5 nm) in a manner inhibited by micromolar ryanodine or ruthenium red. pA (0.5-40 microm) failed to induce SR Ca(2+) release. Rather, pA enhanced Ca(2+) loading into SR and fully inhibited Ca(2+)-, caffeine-, and sMCa-induced Ca(2+) release. The two peptides modified single channel gating behavior in distinct ways. With Cs(+)-carrying current, 10 nm to 1 microm sMCa induced long lived subconductances having 48% of the characteristic full open state and occasional transitions to 29% at either positive or negative holding potentials. In contrast, pA stabilized long lived channel closures with occasional burst transitions to 65% (s1) and 86% (s2) of the full conductance. The actions of pA and sMCa were observed in tandem. sMCa stabilized additional subconductance states proportional to pA-induced subconductances (i.e. 43% of pA-modified s1 and s2 substates), revealing a proportional gating mechanism. [(3)H]Ryanodine binding and surface plasmon resonance analyses indicated that the peptides did not interact by simple competition for a single class of mutually exclusive sites on RyR1 to produce proportional gating. The actions of sMCa were also observed with ryanodine-modified channels and channels deficient in immunophilin 12-kDa FK506-binding protein. These results provide evidence that sMCa and pA stabilize distinct RyR1 channel states through distinct mechanisms that allosterically stabilize gating states having proportional conductance.
authors.affiliation: Department of Molecular Biosciences and Graduate Program in Neurosciences, University of California, Davis, CA 95616, USA.
authors.forename: Lili
authors.initials: L
authors.lastname: Chen
authors.forename: Eric
authors.initials: E
authors.lastname: Estève
authors.forename: Jean-Marc
authors.initials: JM
authors.lastname: Sabatier
authors.forename: Michel
authors.initials: M
authors.lastname: Ronjat
authors.forename: Michel
authors.initials: M
authors.lastname: De Waard
authors.forename: Paul D
authors.initials: PD
authors.lastname: Allen
authors.forename: Isaac N
authors.initials: IN
authors.lastname: Pessah
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D002110:Caffeine / Q000494:pharmacology; D002118:Calcium / Q000378:metabolism; D015640:Ion Channel Gating; D008969:Molecular Sequence Data; D015688:Oncogene Protein pp60(v-src) / Q000494:pharmacology*; D010446:Peptide Fragments / Q000494:pharmacology*; D011817:Rabbits; D012433:Ryanodine / Q000378:metabolism; D019837:Ryanodine Receptor Calcium Release Channel / Q000737:chemistry / Q000187:drug effects* / Q000378:metabolism; D012519:Sarcoplasmic Reticulum / Q000378:metabolism; D012604:Scorpion Venoms / Q000494:pharmacology*
chemical_list: D010446:Peptide Fragments; D019837:Ryanodine Receptor Calcium Release Channel; D012604:Scorpion Venoms; C406406:maurocalcine; C098708:peptide A; D012433:Ryanodine; D002110:Caffeine; D015688:Oncogene Protein pp60(v-src); D002118:Calcium