Fig. 6.

Scoparone facilitated CaM1 binding to the IQ domain of L-VGCC. a GST pull-down assay for CaM1 (K_df, 0.61 μM) binding to peptides (NT, CT1, CT1A, CT1B, PreIQ, and IQ peptides) under 1 mM Ca²⁺. CaM1 binding was examined in the presence of scoparone (EC₅₀, 5.54 μM). BSA (2 μg) and CaM1 (2 μg) were run as standards. b Summary of CaM1 binding by scoparone under 1 mM Ca²⁺. c-e Dose-dependent binding of CaM1 to IQ under 1 mM Ca²⁺ (c, d) and 100 nM Ca²⁺ (c, e) in the presence of 5.54 μM scoparone. For comparison, curves of CaM1 binding in the absence of scoparone were taken from Fig. 5 or constructed, and the fitted curves were superimposed as solid lines. f, g GST fusion peptides (WT-IQ, MT-AA, 0.61 μM) were mixed with 0.61 μM CaM1. h Representative recorded currents and the residual fractions of peak currents (I_res/I_pk) from L-VGCC (WT-IQ) and its mutants (MT-AA) during test pulses of V_h from −80 to + 40 mV. I_Ca was scaled to the peak I_Ba. i Representative recorded currents and I_res/I_pk from L-VGCC (WT-IQ) and its mutants (MT-AA) after pretreatment with 5.54 μM scoparone during test pulses of V_h from −80 to + 40 mV. The I_res/I_pk remaining at the end of a 2-s test pulse was plotted for I_Ca and I_Ba supported by WT-IQ and MT-AA. The data are shown as the mean ± SD from six independent experiments. *p < 0.05, **p < 0.01, ***p < 0.001, ****p < 0.0001. ns, not significant.