Production, Purification and Characterization of Thermostable Protease From Alkaliphilic and Thermophilic Geobacillus Sp.

Abstract

Proteases are the hydrolase enzymes that catalyze the hydrolysis of the peptide bonds in the primary structure of proteins and peptids. They are used to cleave the proteins specifically to produce useful peptides in the processes. Proteases are present in a wide variety of living organisms and they also show different physicological, physicochemical, biological, chemical functions on the earth. They are the most important enzymes in the industry, accounting for 60% of the total enzyme scales in the world. The microorganisms that were previously isolated and characterized as a Bacillus sp. from Balçova Geotermal region in İzmir were used in the experiments. The aim of this study was to produce the protease enzyme from alkaliphilic and thermophilic Bacillus sp., purify and determine the properties of the enzyme with the characterization steps. When the screening studies and growth conditions were investigated, it was understood that the alkaliphilic and thermophilic Bacillus sp. produced extracellular protease enzyme. This extracellular protease enzyme was purified by ammonium sulphate precipitation and ion exchange chromatography chromatograpy. The yield and purification fold after purification of the enzyme were 33% and 1.41, respectively. In the characterization studies, the results indicated that the protease enzyme had highest activity at pH 8.0 and 55 C. The protease enzyme lost 20% of its activity at pH 4.0 and it lost 10% of its activity at pH 10.0. The protease enzyme at temperatures below 55 C lost 15% of its activity and also the protease enzyme at temperatures above 55 C lost 25% of its activity. The protease enzyme was stable at different pH values during 3 hours and at different temperature values during 6 hours. When compared the substrates, casein showed higher activity. The effect of organic solvents and surfactants on protease activity was investigated and the results indicated that the protease enzyme was stable in the presence of 10% of the organic solvents and 1% of the surfactants. PMSF and the protease inhibitor coctail decrease the activity of the protease.