Figure 7.
Binding of the recombinant vf14-3-3s to the phosphorylated H+-ATPase. A, Expression and purification of GST-vf14-3-3 fusion proteins in E. coli. The GST-vf14-3-3s were separated by SDS-PAGE and stained with silver. B, Far western-blot analysis for the H+-ATPase. GCPs were incubated under background RL for 30 min, and were then illuminated with BL for 2.5 min. GCPs at the indicated amounts were subjected to SDS-PAGE, and Far western blot-analysis was conducted using 0.1 μm of GST-vf14-3-3a and b as probes. C, Western-blot analysis for the H+-ATPase. GCPs were loaded as shown in B. D, Relative amount of vf14-3-3s bound to the H+-ATPase. Binding levels were determined densitometrically.