Purification and characterization of thermostable ?-amylase from thermophilic Anoxybacillus flavithermus

Abstract

This study reports on the purification and characterization of thermostable alpha-amylase (alpha-1-4 D-glucan glucanohydrolase EC 3.2.1.1) from a newly isolated Anoxybacillus flavithennus. A. flavithermus was used, which was isolated from hot water springs of Omer, Afyonkarahisar, Turkey. The gram-positive, spore-forming, motile, moderately thermophilic bacteria were found to be a strain of A. flavithermus analysed by 16S rRNA comparison. The optimal conditions for bacterial growth were determined to be at 20th h, 55 degrees C and pH 6.0. Maximum alpha-amylase activity was obtained at 55 degrees C at pH 7.0 after 24h of incubation. Thermostable alpha-amylase from A. flavithermus was purified by 70% (NH4)(2)SO4 and ion-exchange chromatography (5.2-fold; 65.8% yield). The molecular weight of alpha-amylase was 60 kDa, as estimated by sodium dodecyl sulphate-polyacrylamide gel electrophoresis (SDS-PAGE). The alpha-amylase hydrolyzed soluble starch at 55 degrees C with K-m: 0.005 mM and V-max: 3.5 mu mol min(-1). (C) 2013 Elsevier Ltd. All rights reserved.