Purification of Amylase Produced by Thermophilic Bacillus sp. TS9

Abstract

Amylases are the starch degrading enzymes obtained from diverse sources such as plants, animals and microorganisms including bacteria and fungi. But the microbial amylases are preferred over other sources. The thermophilic alpha amylase obtained from bacterial species has applications in many industries. Six thermophilic bacteria Microbacterium sp. TS1, Pseudomonas sp. TS2, Alishewanella sp.TS3, Rheinheimera sp. TS5, Bacillus sp. TS9 and halophilic Bacillus sp. HS7 were isolated from Shahdrah and Kheiwera regions of Pakistan and their 16s RNA identified through molecular techniques. All the isolates were screened for amylase quantitatively and qualitatively. The Thermophilic Bacillus sp. TS9, based on its higher production of amylase, was selected for purification and characterization. The bacteria produced clearance zone of 2.7cm in diameter on 1% starch supplemented agar plate and the maximum specific activity of 2.45U/mg was obtained after 72 hours incubation at 45° C and pH 8. The extracellular amylase was then purified by ammonium sulphate precipitation and gel permeation chromatography. The specific activity of amylase obtained from Bacillus sp. TS9 increased to 4.47U/mg (1.82 fold) through partial purification, and 15.92U/mg (6.48 fold) through column chromatography. The current Study points out a valuable approach to purifiy the amylase and consequently improves the activity of amylase which have numerous applications in many industries