Visualization of leucineaminopeptidase activity after acrylamide gel electrophoresis

Abstract

Several methods were described for visualization of proteolytic activity on electrophoregrams obtained with agar, agarose, starch, or acrylamide gels as supporting media. In most of these reports casein or hemoglobin were used as nonspecific substrates (1–3). Recently, colorimetric assays for trypsin, using α-benzoyl-d,l-arginine-p-nitroanilide (4), and for subtilisin, using Z-glycyl-glycyl-l-leucine-p-nitroanilide (5) were introduced for the localization of these proteases after acetate celulose and acrylamide gel electrophoresis. No convenient and simple methods were in practice for detection of leucineaminopeptidase (3), although this enzyme was assayed in solution with specific chromogenic substrates such as l-leucine-p-nitroanilide or l-leucine-β-naphtylamide (6,7).The present report describes the use of p-nitroanilide substrate-l-leucine-p-nitroanilide for detection of leucineaminopeptidase activity after acrylamide gel electrophoresis. The method allows a rapid and sensitive localization of leucineaminopeptidases.