PURIFICATION AND CHARACTERIZATION OF THERMOSTABLE ALPHA-AMYLASE-II FROM BACILLUS-SP-JF(2) STRAIN

Thermostable alpha-amylase from Bacillus sp-JF(2) strain was found to have three ee active components (named alpha-amylase I, II, and III) with molecular weights of 110,000, 140,000, and 300,000, respectively. alpha-Amylase II was isolated and purified in the current work by different procedures from that for alpha-amylase I. alpha-Amylase II consists of two identical subunits (MW 70,000). The isoelectric point is 4.7. The temperature optimum is at 90 degrees C and the pH optimum is 5.5 for the enzyme activity. The half-life of the enzyme at 90 degrees C is 30 min, and the enzyme is stable over a pH range of 7.0-9.0. The Km value of the enzyme was estimated to be 3.3 mg ml(-1). A considerable difference in amino acid composition was observed between alpha-amylase I and alpha-amylase II. The alpha-helix content of alpha-amylase II was calculated to be 51% from the circular dichroism spectrum. The number of Ca2+ binding to each molecule of alpha-amylase II was determined to be 10 by atomic absorption.