Isoelectric focusing, effect of reducing agents and inhibitors: partial characterization of proteases extracted from Bromelia karatas
Tóm tắt
The aim of this research is the partial characterization of proteases extracted from B. karatas; the isolation and purification of proteases from B. karatas fruits were achieved using precipitation, separation by size exclusion chromatography and anion-exchange chromatography; molecular mass (MM) was determined, and the effect of inhibitors, reducing agents and heat on enzyme activity was analyzed. These proteases were compared with proteases from Bromelia pinguin (B. pinguin) and evaluated under similar conditions. The isolation procedure was adequate; only a few protein bands are present in sodium dodecyl sulfate polyacrylamide gel electrophoresis. Furthermore, zymogram analysis showed protein bands with enzyme activity. Inhibitors, reducing agents and heat were unable to inactivate the proteases extracted from B. karatas and B. pinguin. The semi-purified extracts are a set of proteases with a MM of 66 kDa, but different isoelectric points (3.5–6.5 for B. karatas and 5–9 for B. pinguin), which are found in quaternary structures with proteolytic activity. When denatured, they segment into fragments of approximately 20 and 10 kDa. The data indicate that these plants could be used as sources of proteases since they present good proteolytic activity (21.93 UT for proteases from B. karatas and 43.58 UT for proteases from B. pinguin) and that B. Karatas has potential applications comparable to B. pinguin in the food and health industries.
Tài liệu tham khảo
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