Gene cloning and characterization of a thermostable organic-tolerant α-amylase from Bacillus subtilis DR8806.

The gene encoding an extracellular α-amylase from Bacillus subtilis DR8806 was cloned into pET28a(+) vector and expressed in Escherichia coli BL21 (DE3). The recombinant enzyme with molecular mass of 76 kDa exhibited optimal activity at pH 5.0 and 70 °C with high stability in pH and temperature ranges of 4.0-9.0 and 45-75 °C. The enzyme showed a half-life of 125 min at 70 °C. The α-amylase activity enhanced in the presence of Na(+), K(+), and Ca(2+) ions, while Zn(2+), Pb(2+), and Hg(2+) ions inhibited the activity. The recombinant α-amylase exhibited high stability towards ioninc detergents sodium dodecyl sulfate (SDS) and cetyl trimethylammonium bromide (CTAB). Organic solvents in reaction media increased the α-amylase activity. TLC analysis showed that maltoriose and maltose were the major end products of enzymatic starch hydrolysis. Presenting various properties of recombinant α-amylase makes it well suited as a potential candidate for industrial usages.

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