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. 2007 Jul 13;405(Pt 3):489–494. doi: 10.1042/BJ20070174

Figure 5. Complex formation between His–centerin and a panel of proteases.

Figure 5

His–centerin was incubated with a panel of proteases at either 1:1 or 5:1 molar ratio (centerin/enzyme) for 30 min at 37 °C and then electrophoresed on SDS/PAGE and Western blotted. His–centerin was detected using anti-His monoclonal antibody and goat anti-mouse HRP-conjugated secondary antibody. (A) The Figure shows the results of incubations of His–centerin with trypsin-like proteases: lane 1, His–centerin with no protease; lane 2, trypsin 1:1; lane 3, trypsin 5:1; lane 4, thrombin 1:1; lane 5, thrombin 5:1; lane 6, plasmin 1:1; lane 7, plasmin 5:1; lane 8, Factor Xa 1:1; lane 9, Factor Xa 5:1. (B) The Figure shows the results of incubations of His–centerin with other serine proteases: lane 1, His–centerin with no protease; lane 2, chymotrypsin 1:1; lane 3, chymotrypsin 5:1; lane 4, cathepsin G 1:1; lane 5, cathepsin G 5:1; lane 6, elastase 1:1; lane 7, elastase 5:1. (C) The Figure shows results obtained using cysteine proteases. Lanes 1 and 2 show a positive control using the serpin MENT, which forms an SDS-stable complex under the conditions of this assay: lane 1, MENT-cathepsin L; lane 2, MENT alone. Lanes 3–7 show the results of incubations of His–centerin with cathepsins L and V at either 1:1 or 5:1 molar ratio (serpin/protease). Lane 3, cathepsin L 1:1; lane 4, cathepsin L 5:1; lane 5, cathepsin V 1:1; lane 6, cathepsin V 5:1; lane 7, His–centerin with no protease.