Partial Purification and Characterization of Alkaline Thermostable Proteases of Jack Mackerel (Trachurus murphyi)

Abstract

The objective of this research was to carry out the extraction, partial purification, characterization and inhibition of the alkaline thermostable proteases of jack mackerel white muscle (Trachurus murphyi). It was determined that the better extraction medium of proteolyt-ic enzymes was 60 mM bicarbonate buffer pH 9.5. The partial purification of protease was carried out applying a thermal shock at 50 °C for 10 min and gel filtration chromatography. Two peaks were collected with proteolytic activity and specific activity of 13.70 and 0.33 U'/mg, respectively. Frozen storage at −18 °C increased proteolytic activity of the extracts from 0.53 U′/ml to 7.1 U′/ml after 161 h. Inactivation constants ranged between 4.65 ×10−3 and 10.78 ×10−3 min−1 for a temperature of 40-65°C. The activation energy of denaturation was 31.85 KJ/ mol, the transition temperature and enthalpy were 59.01°C and 1.16 J/g, respectively. The protease was partially inactivated with 1.0 mM phenylmethylsulfonyl fluoride (PMSF) and 1 mM iodoacetamide (IA).