Fig 1: Proteolytic activity of L. interrogans leptolysin on proteoglycans and plasma fibronectin. (A) Degradation of substrates: mimecan, lumican, biglycan, decorin (0.5 μg), and plasma fibronectin (5 μg) were incubated with leptolysin (0.1 µg) at 37°C for the indicated time points. Cleavage products were subjected to SDS- polyacrylamide gel under reducing conditions, transferred to nitrocellulose membranes, and probed with specific antibodies, or the gel was silver stained (FP). (B) Inhibition of proteolytic activity: before the addition of each substrate, leptolysin (0.1 µg) was incubated with 5 mmol/L 1,10-phenanthroline (lane 4) for 30 min at room temperature. Substrates were added and incubations proceeded for 24 h. Cleavage products were analyzed as described above. Mim (mimecan), Lum (lumican), Bgn (biglycan), Dec (decorin), FP (plasma fibronectin), Sup (Leptospira culture supernatant), Leptol. (leptolysin), Fen (1,10-phenanthroline).