Figure 8

SspA degrades occludin and collagen IV. A pcDNA-occludin-myc (2.5 µg) was co-transfected with pcMV-sspA-flag (0, 0.5, 1.0, 1.5, 2.0, or 2.5 µg; corresponding to Lanes 1–6). The occludin level was measured by Western blotting, and data were analysed using the Quantity One program. The results showed that with increasing expression of SspA, the expression of occludin decreased gradually. B Occludin was incubated with the secreted proteins of Yb2, Yb2ΔsspA, or cYb2ΔsspA or with recombinant SspA at an enzyme:substrate weight ratio of 1:1000 at 37 °C in buffer (100 mM Tris, 150 mM NaCl, 5 mM CaCl₂, 0.02% NaN3; pH 8.0). At specific time points, aliquots were collected for Western blotting and data analysis using the Quantity One program. Occludin incubated with ADCF-mAb medium was used as the control. C SspA degrades collagen IV. Collagen IV was incubated with the secreted proteins of Yb2, Yb2ΔsspA, or cYb2ΔsspA or with recombinant SspA at an enzyme:substrate weight ratio of 1:1000 at 37 °C in buffer (100 mM Tris, 150 mM NaCl, 5 mM CaCl₂, 0.02% NaN₃; pH 8.0). At specific time points, aliquots were collected for Western blotting and data analysis using the Quantity One program. Collagen IV incubated with ADCF-mAb medium was used as the control. The data are presented as the means ± standard deviations of the results of three independent experiments conducted in triplicate. Statistical significance was assessed using one-way or two-way analysis of variance (ANOVA). The asterisks indicate the level of statistical significance (***, p < 0.001; ****, p < 0.0001). ns: not significant.