Figure 4.

Fluorescence imaging of an SDS/PAGE showing the results of proteolytic digestions of Csk^PEP and Csk^pPEP with subtilisin. Lane 1: Csk^pPEP − subtilisin; lane 2: Csk^pPEP + subtilisin; lane 3: Csk^PEP − subtilisin; lane 4: Csk^PEP + subtilisin. Reactions conditions: Csk^PEP and Csk^pPEP (1 μg) in 20 μl buffer (20 mM Tris⋅acetate, pH 8.0/10% glycerol/2 mM DTT) treated with subtilisin Carlsberg (12.5 ng) for 30 min at 4°C. Fluorescence imaging was done on a Storm instrument (Molecular Dynamics). Proteolysis reactions were performed in parallel under identical conditions with different protein preparations on four different occasions and all gave comparable results to those shown here.