Enzymatic activity of proteases from Cyprinus carpio (Cypriniformes: Cyprinidae) captured in a polluted lagoon in Mexico
Common carp (Cyprinus carpio) is an aquatic organism of commercial value able to survive in polluted environments; carps contain proteolytic enzymes of physiological importance and potential industrial application. The objective of this work was partially purify and study the proteolytic activity at different pH of carp proteases living in a polluted environment. Three carps were captured in different zones of Zumpango polluted lagoon (Mexico) at 1 m of maximum deep. Protease crude extracts were obtained from dorsal muscle by aqueous extraction and fractionated by 20 %, 50 %, 80 %-saturated (NH4)2SO4. Fractions extracted with 50 % and 80 %-saturated (NH4)2SO4 were selected for their high proteolytic activity and concentrated by ultrafiltration through 100 kDa molecular weight cutoff membranes and analyzed by sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE). The crude proteolytic extract had significantly higher activity (19.7 - 20.3 U / mg) at pH 2, 5, and 7 (P < 0.001). Fractions obtained with 20 %, 50 % and 80 % - saturated (NH4)2SO4 showed peak activity at pH 5 (2.8 U / mg) and pH 6 (2.2 U / mg); pH 6 (4.3 U / mg) and pH 3 - 4 (3.6 - 3.7 U / mg); pH 3 (10.8 U / mg) and pH 10 (10.6 U / mg); respectively. Subfractions of < 100 kDa, obtained with 50 % and 80 %-saturated (NH4)2SO4, had peak proteolytic activity at alkaline pH. A < 100 kDa fraction, obtained with 80 %-saturated (NH4)2SO4, had the highest proteolytic activity (37.3 - 43.7 U / mg) at pH 8 - 10, purification factor of 3 and 19.1 % recovery. Thirteen proteins between 9.8 to 104.8 kDa were identified in the crude extract. Peak protein concentration was observed for 31 - 33 and 39 - 41 kDa, suggesting the possibility predominance of serine- and aspartyl- proteases, respectively. We suggest this protease with maximum activity at alkaline pH is related to the adaptation of C. carpio to polluted waters with high pH. Although unsuitable for human consumption, these organisms can be a source of protease production aimed to several uses as in the industry and waste water treatment among others.
Keywords: common carp, Cyprinus carpio, proteases, contaminated water, lagoon