Figure 1. TnT digestion by calpain under exhaustive conditions.

TnT (1.1 μg) was incubated with 100 mU of calpain-1 (C1) or -2 (C2) in the presence of 5 mM CaCl₂ at 30°C for the indicated time (left panel) or 20 min (right panel). Lane 1, TnT; lanes 2, 8, 9, C1; lanes 3, 4, 10, 11, TnT and C1; lanes 5, 6, 12, 13, TnT and C2; lane 7,14, 15, C2. For lanes 8, 10, 13 and 15, incubation was carried out with additional 50 mM EDTA. The gel was run with 2× running buffer [23] for better resolution of low molecular weight fragments (right panel). Open and black arrowheads, intact TnT and calpain catalytic subunit, respectively; arrow, calpain small subunit; *, proteolyzed TnT fragments; bracket, autolytic fragments of calpain.