Figure 4.

N-terminus of hSCGN is essential for calcium response. (A) Purified hSCGN WT and ΔEF1 protein were pre-treated with 0, 2 mM CaCl₂ for 15 min at R.T. and then incubated with the indicated concentrations of H₂O₂ for 1 h at 37°C. Coomassie blue-stained SDS-PAGE gels under non-reducing (−β-ME) and reducing (+β-ME) condition. (B) HeLa cells overexpressing hSCGN WT and ΔEF1 were treated with 0 or 2 μM ionomycin for 5 min and with 0 or 5 mM H₂O₂ for 30 min at 37 °C. Cell lysates were subjected to Western analysis using anti-hSCGN or anti-α-tubulin antibody. Non-specific bands were indicated as *. (C) Recombinant hSCGN WT and ΔEF1 proteins were incubated with 0 or 2 mM CaCl₂ for 15 min, and with 1 mM NPSB-B for 2 h at R.T. Labeled samples in gel sample buffer containing 10 mM NEM were detected with streptavidin-HRP. Full-length gels and blots are in Supplementary Fig. S7.