Purification and characterization of novel ?-amylase from Bacillus subtilis KIBGE HAS.

ABSTRACT: Purification of extracellular ?-amylase from Bacillus subtilis KIBGE HAS was carried out by ultrafiltration, ammonium sulfate precipitation and gel filtration chromatography. The enzyme was purified to homogeneity with 96.3-fold purification with specific activity of 13011 U/mg. The molecular weight of purified ?-amylase was found to be 56,000 Da by SDS-PAGE. Characteristics of extracellular ?-amylase showed that the enzyme had a Km and V (max) value of 2.68 mg/ml and 1773 U/ml, respectively. The optimum activity was observed at pH 7.5 in 0.1 M phosphate buffer at 50 °C. The amino acid composition of the enzyme showed that the enzyme is rich in neutral/non polar amino acids and less in acidic/polar and basic amino acids. The N-terminal protein sequence of 10 residues was found to be as Ser-Ser-Asn-Lys-Leu-Thr-Thr-Ser-Trp-Gly (S-S-N-K-L-T-T-S-W-G). Furthermore, the protein was not N-terminally blocked. The sequence of ?-amylase from B. subtilis KIBGE HAS was a novel sequence and showed no homology to other reported ?-amylases from Bacillus strain.

SUBMITTER: Bano S 

PROVIDER: S-EPMC3066380 | biostudies-literature | 2011 Mar

REPOSITORIES: biostudies-literature