Detection of exo-beta-1,3-glucanase activity in polyacrylamide gels after electrophoresis under denaturing or nondenaturing conditions.
A method for the visualization of exo-beta-1,3-glucanase activity in polyacrylamide gels is presented. The procedure consists of the enzyme reaction in the gel with the substrate alpha-naphthylglucopyranoside, and a subsequent staining of the obtained alpha-naphthol with dyes Fast Red B, or Fast Blue BB, respectively. A mixture of exoglucanases produced by the fungus Polyporus squamosus was used for the optimization of the method. The procedure is applicable for the standard Laemmli discontinuous electrophoresis system, even in the presence of sodium dodecyl sulfate, as well as for electrophoresis in linear gradients of the polyacrylamide concentration. The staining method was used for the analysis of exoglucanases secreted by several yeast genera. All yeasts tested produced two types of exoglucanases, a high molecular mass species heterogeneous in size, and one or two smaller homogeneous enzymes.
Vargić
T
T
Faculty of Food Technology and Biotechnology, University of Zagreb, Croatia.
Mrsa
V
V
D003720:Densitometry; D004591:Electrophoresis, Polyacrylamide Gel / Q000191:economics / Q000379:methods*; D005656:Fungal Proteins / Q000032:analysis*; D043326:Glucan 1,3-beta-Glucosidase; D011489:Protein Denaturation; D012441:Saccharomyces cerevisiae / Q000201:enzymology; D012680:Sensitivity and Specificity; D013194:Staining and Labeling; D013997:Time Factors; D001617:beta-Glucosidase / Q000032:analysis*
D005656:Fungal Proteins; D001617:beta-Glucosidase; D043326:Glucan 1,3-beta-Glucosidase