Purification, characterization of thermostable Amylopullulanase from Bacillus licheniformis (BS18) by using solid state fermentation (SSF)

Abstract

An amylopullulanase enzyme has a wide range of applications in the food processing and distillery industries, including the conversion of starch to sugers and the production of conversion syrups (maltose and fructose syrups). Amylopullulanase was purified by three stepsincluded precipitation with 40% saturation ammonium sulfate, ion exchange chromatography by DEAE- cellulose column, and gel filtration by Sephacryl S-200 column. The characteristics of purified amylopullulanase were studied. The optimum pH of enzyme activity was 7.0. It was more stable at pH 7, andthe maximum enzyme activity was observed at 70°C. The optimum thermal stability for the enzyme was at (37-50°C). The effect of some metal ions on amylopullulanase activity was determined. It was observed that 10 mM of Ca++ and Mg++ enhanced enzymes activity. Different levels of inhibition revealed when enzyme was treated with Fe++, Cu++, Zn++, and Hg++ at 5 and 10 mM. An effect of some chemical agents on purified enzyme activity was investigated. Results obtained that a slight effect the amylopullulanase activity occurred after incubation with 5, 10 of EDTA and 10mM of PMSF on amylase activity. Hence, amylopullulanase activity have increased effect after incubation with 5, 10mM of 2-mercaptoethanol, while amylopullulanase lost most of its activity after incubation with 5M of urea.