Figure 7
CSQ1 ∆C14 binds to and activates purified RyR1. (A) Records of 3 s of single channel activity, where channel opening is upward from zero current (c, continuous line) to maximum open conductance (o, broken line) at +40 mV. (A) Control purified RyR1 activity, with 2 mM cis ATP and 100 nM cis Ca2+ free (top trace) and after the addition of 16 μg/ml CSQ1∆C14. (B) Average relative data (relative to activity before the addition of WT CSQ1 or CSQ1∆C14; N = 9) showing open probability (P o ). Average data significantly different (P ≤ 0.05) from channel activity recorded in the absence of CSQ1’s is indicated by asterisk (*). Crosshatch (#) indicates a significant difference (P ≤ 0.05) between the average relative P o recorded in the presence of WT CSQ1 and CSQ1∆C14. (C) CSQ1 affinity chromotography. WT CSQ1 (left) and CSQ1∆C14 (right) after exposure to purified RyR1. Binding was repeated three times. Blot was immunoprobed with antibodies against RyR1 (top) and CSQ1 (bottom). (D). Purification of RyR1 from SR vesicles. Immunoprobing purified RyR1 sample with anti-CSQ1, anti-trisk95, and anti-junctin shows no contaminant levels of these proteins in the purified sample. Molecular weight marker is between blots in (C) and (D).