Figure 4B The time course of the distance between the geometrical centers of domains a and a′ and the angle among domains b, b′ and a′ from the simulation of pS357-PDI (red) and PDI (black) (Yang et al., 2014).C (Left) ANS fluorescence spectra of PDI WT and ­­S357E. Controls with ANS alone was shown. A.U., arbitrary units. (Right) Enhancement factor were quantified. Data information: All data were shown as mean ± SEM of three independent experiments. **p < 0.01, ***p < 0.001 (two-tailed Student's t-test).D Limited proteolysis of PDI WT (Left) and S357E (Right) proteins by proteinase K (Pro K) for indicated times. Main digested fragments were indicated on the right margin. Band abb'x* is a digested product with cleavage after Met356 in the x-linker region (Wang et al., 2010).E shCtrl and shFAM20C HepG2 cells were treated without or with 5 μM Tg for 30 min, and in some aliquots of cells Tg was removed for 30 min. Cell lysates were subjected to proteinase K digestion for 3 min and immunoblotting analysis.F HepG2 cells expressing HA-tagged PDI and FLAG-tagged Fam20C with different ratios of plasmid concentrations were subjected to proteinase K digestion for 21 min and immunoblotting analysis.Chaperone activities of PDI WT and S357E (G, H) or phosphorylated PDI WT and S357A by Fam20C for indicated times (I Denatured and reduced GAPDH (G, I) and rhodanese (H, were used as substrates, respectively. The chaperone activity of PDI WT was taken as 100%. Data information: All data were shown as mean ± SEM of three independent experiments. , **p < 0.01, ***p < 0.001 (two-tailed Student's t-test). In (I , ***p < 0.001 (two-way ANOVA, the post hoc Tukey's HSD test).Chaperone activities of phosphorylated PDI WT and S357A by Fam20C for indicated times J). Denatured and reduced rhodanese J) were used as substrates, respectively. The chaperone activity of PDI WT was taken as 100%. Data information: All data were shown as mean ± SEM of three independent experiments. ***p < 0.001 (two-way ANOVA, the post hoc Tukey's HSD test).Reductase activities of PDI WT and S357E (K) measured by insulin reduction assay. The activity of PDI WT was taken as 100%. Data information: All data were shown as mean ± SEM of three independent experiments. **p < 0.01, ***p < 0.001 (two-tailed Student's t-test).Isomerase activities of PDI WT and S357E (L) measured by scrambled RNase A reactivation assay. The activity of PDI WT was taken as 100%. Data information: All data were shown as mean ± SEM of three independent experiments. , **p < 0.01, ***p < 0.001 (two-tailed Student's t-test).Reductase activities of phosphorylated PDI WT and S357A by Fam20C for indicated times (M) measured by insulin reduction assay. The activity of PDI WT was taken as 100%. Data information: All data were shown as mean ± SEM of three independent experiments. , ***p < 0.001 (two-way ANOVA, the post hoc Tukey's HSD test).Isomerase activities of phosphorylated PDI WT and S357A by Fam20C for indicated times (N) measured by scrambled RNase A reactivation assay. The activity of PDI WT was taken as 100%. Data information: All data were shown as mean ± SEM of three independent experiments. , ***p < 0.001 (two-way ANOVA, the post hoc Tukey's HSD test).