| Article ID | Journal | Published Year | Pages | File Type |
|---|---|---|---|---|
| 18369 | Enzyme and Microbial Technology | 2007 | 5 Pages |
Crude extracts were partially purified by organic solvents fractionation: best results (96% of proteins, 91% of total caseinolytic activity) were obtained by adding four volumes of cold acetone to the crude extract. This preparation (redissolved acetone precipitate, RAP) showed maximum activity (>80%) at pH 5–9, and exhibited high thermal stability (>90% of residual activity after heating for 60 min at 60 °C). The enzyme was completely inhibited by E-64 trans-epoxysuccinyl-leucyl-amido(4-guanidino)-butane and iodoacetic acid and activated by the addition of cysteine or β-mercaptoethanol; these results strongly suggest that the isolated protease should be included within the cysteine group, as all the other studied proteases belonging to the family Bromeliaceae. IEF-zymogram of RAP showed five bands (pI 7.3 to <9.3), most of them proteolytically actives, but only three of which (pI 7.6, 8.2 and 8.8) proved to be important. Ion exchange chromatography in DEAE-Sephadex, was selected to separate this active bands.
