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Purification, characterization and substrate specificity of a trypsin from the Amazonian fish tambaqui (Colossoma macropomum)

2010; Elsevier BV; Volume: 396; Issue: 3 Linguagem: Inglês

10.1016/j.bbrc.2010.04.155

1090-2104

Marina Marcuschi, Talita S. Espósito, Maurício F.M. Machado, Izaura Yoshico Hirata, Marcelo F. Marcondes, Márcia Vanusa da Silva, Luiz Bezerra de Carvalho, Vitor Oliveira, Ranilson de Souza Bezerra,

Protein Hydrolysis and Bioactive Peptides

An enzyme was purified from the pyloric caecum of tambaqui (Colossoma macropomum) through heat treatment, ammonium sulfate fractionation, Sephadex G-75 and p-aminobenzamidine-agarose affinity chromatography. The enzyme had a molecular mass of 23.9 kDa, NH(2)-terminal amino acid sequence of IVGGYECKAHSQPHVSLNI and substrate specificity for arginine at P1, efficiently hydrolizing substrates with leucine and lysine at P2 and serine and arginine at P1'. Using the substrate z-FR-MCA, the enzyme exhibited greatest activity at pH 9.0 and 50 degrees C, whereas, with BAPNA activity was higher in a pH range of 7.5-11.5 and at 70 degrees C. Moreover, the enzyme maintained ca. 60% of its activity after incubated for 3h at 60 degrees C. The enzymatic activity significantly decreased in the presence of TLCK, benzamidine (trypsin inhibitors) and PMSF (serine protease inhibitor). This source of trypsin may be an attractive alternative for the detergent and food industry.