A new method for the detection of proteolytic activity in Pseudomonas lundensis after sodium dodecyl sulfate-polyacrylamide gel electrophoresis

Abstract

A new method for the visualization of proteolytic activity in cell culture supernatant from Pseudomonas lundensis after sodium dodecyl sulfate (SDS)--gel electrophoresis is described. Following conventional electrophoresis, the gel is washed in a methanol-containing buffer to facilitate partial removal of SDS. After incubation with 0.5% casein the gel is stained for protein with Coomassie Brilliant Blue R-250. Bands with proteolytic activity appear as clear areas in the gel against a blue-stained background. Molecular weight standards electrophoresed in the same gel stain more intensely than the background and allow determination of the molecular weights of the proteolytic components. The sensitivity of post-electrophoretic reactivation in SDS-gels was determined using trypsin as standard. A slight modification of the technique allowed detection of proteolytic activity in nondenaturing and in isoelectric focusing gels.