Suramin and suramin analogs activate skeletal muscle ryanodine receptor via a calmodulin binding site

Contraction of skeletal muscle is triggered by the rapid release of Ca2+ fr om the sarcoplasmic reticulum via the ryanodine receptor/calcium-release ch annel. The trypanocidal drug suramin is an efficient activator of the ryano dine receptor, Here, we used high-affinity [H-3]ryanodine binding to sarcop lasmic reticulum from rabbit skeletal muscle to screen for more potent anal ogs of suramin. This approach resulted in the identification of NF307, whic h accelerates the association rate of [H-3]ryanodine binding with an EC50 = 91 +/- 7 mu M at 0.19 mu M calculated free Ca2+. In single-channel recordi ngs with the purified ryanodine receptor, NF307 increased mean open probabi lity at 0.6 mu M Ca2+ from 0.020 +/- 0.006 to 0.53 +/- 0.07 with no effect on current amplitude and unitary conductance. Like caffeine, NF307 exerts a very pronounced Ca2+-sensitizing effect (EC50 of Ca2+ shifted similar to 1 0-fold by saturating NF307 concentrations). Conversely, increasing concentr ations of free Ca2+ sensitized the receptor for NF307 (EC50 = 146 +/- 3.5 m u M at 0.82 mu M estimated free Ca2+). The effects of NF307 and caffeine on [H-3]ryanodine binding were additive, irrespective of the Ca2+ concentrati on. In contrast, the effects of calmodulin, which activates and inhibits th e ryanodine receptor in the absence and presence of Ca2+, respectively, and of NF307 were mutually antagonistic. If the purified ryanodine receptor wa s prebound to a calmodulin-Sepharose matrix, 100 mu M NF307 and 300 mu M su ramin eluted the purified ryanodine receptor to an extent that was comparab le to the effect of 10 mu M calmodulin, We conclude that NF307 and suramin interact directly with a calmodulin binding domain of the ryanodine recepto r. Because of its potent calcium-sensitizing effect, NF307 may represent a lead compound in the search of synthetic ryanodine receptor ligands.