Figure 3. FKBP12 dissociation from SR vesicles in the presence of NOC12.

(A–C) SR vesicles not treated with FK506 were incubated for 5 h at 24°C with or without 0.10 mM NOC12 in the absence and presence of 44 µM S107 in 0.25 M KCl, 20 mM imidazole, pH 7.0, 7 µM free Ca²⁺ and protease inhibitors. S-nitrosylation was stopped by centrifugation. Resuspended samples were separated on 8–20% (FKBP12) and 3–12% (RyR1 and Cys-SNO) gradient SDS-PAGE gels and transferred to nitrocellulose membranes to detect S-nitrosylation of RyR1, and FKBP12 and RyR1 proteins. Data are the mean ± SEM of 4 determinations. *p<0.05 compared to control samples (B) and samples with NOC12 and S107 (C). (D and E) SR membranes were incubated with and without 44 µM S107 and 0.1 mM NOC12 at 24°C for 90 min, solubilized, and immunoprecipitated as described in Methods. Immunoblots of RyR1 and FKBP12 are shown. Data are the mean ± SEM of 4 experiments. ^*p<0.05 compared to control samples and samples incubated with NOC12 and S107.