ABSTRACT Commercially grown Crimini mushrooms were extracted in the presence of a variety of serine, cysteine, acidic, and metallo protease inhibitors to examine the effect of these inhibitors on the recovery of tyrosinase isoforms. Native PAGE showed similar types of tyrosinase isoforms, but only extracts containing EDTA and a fungal inhibitor cocktail showed new forms. SDS PAGE showed only minor differences in the total protein profiles except for extracts containing the fungal inhibitor cocktail. Western blotting identified a 45-48 kD tyrosinase in all extracts, but some extracts contained a 64 kD latent tyrosinase. Isoelectric focusing showed a total of at least 7-8 tyrosinase isoforms, however, the pattern and distribution of isoforms seemed to vary according to the type of protease inhibitor used in the initial extraction. Treatment of control extracts with proteases showed that subtilisin was more effective in increasing the activity of 5.0 and 5.6 pI isoforms. Protease inhibitors do not seem to preserve latency of tyrosinase or change the isoforms composition significantly, suggesting that post-harvest handling/processing of Crimini mushrooms at this stage has already resulted in the activation of tyrosinase and change in isoform composition.
Buy this Article
|