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The current paper reports on the production and characterization of extracellular thermostable proteases produced from thermophilic bacteria namely Aeribacillus pallidus VP3, Anoxybacillus kamchatkensis M1V, Virgibacillus natechei FarDT, and Melghiribacillus thermohalophilus Nari2AT strains isolated from different extremophlic biotopes in Tunisia and Algeria. The key challenge of the use of the different proteases in industrial applications is their efficiency under unconventional conditions. An optimization via one-factor-at-a-time (OFAT) methodology has been used to obtain 3,000 U.mL-1; 4,600 U.mL-1; 15,800 U.mL-1; and 16,000 U.mL-1 of proteases activity from VP3, M1V, FarDT, and Nari2AT strains, respectively. Particularly, VP3 and FarDT proteases showed a high tolerance to several organic solvents. In addition, the protease produced from FarDT strain might be useful as potential candidate for application in detergency. The deproteinization of shrimp wastes by M1V and Nari2AT proteases was successful with a high rate of deproteinization of about 48% and 82%. The obtained proteolytic hydrolysate obtained by M1V strain showed high biological activities with half maximal inhibitory concentration (IC50) of 71.52 µg.mL-1, 107.67 µg.mL-1, and 133.24 µg.mL-1, respectively for the angiotensin, tyrosinase, and amylase inhibitory activities. [ABSTRACT FROM AUTHOR]

The present study investigates the purification and biochemical characterization of a novel extracellular serine alkaline protease, subtilisin (called SAPN) from Melghiribacillus thermohalophilus Nari2AT. The highest yield of protease (395 IU/g) with white shrimp shell by-product (40 g/L) as a unique source of nutriments in the growth medium was achieved after 52 h at 55 °C. The monomeric enzyme of about 30 kDa was purified to homogeneity by ammonium sulfate fractionation, heat treatment, followed by sequential column chromatographies. The optimum pH and temperature values for subtilisin activity were pH 10 and 75 °C, respectively, and half lives of 9 and 5 h at 80 and 90 °C, respectively. The sequence of the 25 NH2-terminal residues pertaining of SAPN exhibited a high homology with those of Bacillus subtilisins. The inhibition by DFP and PMSF indicates that this enzyme belongs to the serine proteases family. SAPN was found to be effective in the deproteinization (DDP %) of blue swimming crab (Portunus segnis) and white shrimp (Metapenaeus monoceros) by-products, with a degree of 65 and 82%, respectively. The commercial and the two chitins obtained in this work showed a similar peak pattern in Fourier-Transform Infrared (FTIR) analysis, suggesting that SAPN is suitable for the bio-production of chitin from shell by-products. Article Highlights: The purification of subtilisin (SAPN) from M. thermohalophilus Nari2A was carried out. The molecular weight and the NH2-terminal sequence of the subtilisin were determined. Optimum pH and temperature values for activity were pH 10 and 75 °C respectively. SAPN was found to be effective in the deproteinization of crab and shrimp by-products. SAPN may be used as candidate for chitin extraction from crustacean by-products. [ABSTRACT FROM AUTHOR]

The present study investigates the purification and biochemical characterization of a novel extracellular serine alkaline protease, subtilisin (called SAPN) from Melghiribacillus thermohalophilus Nari2A T . The highest yield of protease (395 IU/g) with white shrimp shell by-product (40 g/L) as a unique source of nutriments in the growth medium was achieved after 52 h at 55 °C. The monomeric enzyme of about 30 kDa was purified to homogeneity by ammonium sulfate fractionation, heat treatment, followed by sequential column chromatographies. The optimum pH and temperature values for subtilisin activity were pH 10 and 75 °C, respectively, and half lives of 9 and 5 h at 80 and 90 °C, respectively. The sequence of the 25 NH 2 -terminal residues pertaining of SAPN exhibited a high homology with those of Bacillus subtilisins. The inhibition by DFP and PMSF indicates that this enzyme belongs to the serine proteases family. SAPN was found to be effective in the deproteinization (DDP %) of blue swimming crab (Portunus segnis) and white shrimp (Metapenaeus monoceros) by-products, with a degree of 65 and 82%, respectively. The commercial and the two chitins obtained in this work showed a similar peak pattern in Fourier-Transform Infrared (FTIR) analysis, suggesting that SAPN is suitable for the bio-production of chitin from shell by-products.