| Summary: | Culture supernatants of ΔPa PRO1 , ΔPa PRO2 , and wild-type GB-4(0) were centrifuged at 20, 000 × g for 5 min, and subjected to SDS-PAGE containing 0.1% gelatin without heat denaturation. After electrophoresis, to remove SDS [ 35 ], the polyacrylamide gel was incubated with 2.5% tritonX-100 at 25°C for 1h, washed with distilled water, and then incubated with buffer fluids, i.e, 0.1M sodium acetate (pH5.2), 0.1M Tris-HCl (pH7.2), or 0.1M Tris-HCl (pH9.8). The gel was stained with Symply Blue Safe Stain (Invitrogen, CA). Protease activity was visualized as white band caused by gelatin degradation. (TIF)
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