A highly thermoactive and salt-tolerant [alpha]-amylase isolated from a pilot-plant biogas reactor

Aiming at the isolation of novel enzymes from previously uncultured thermophilic microorganisms, a metagenome library was constructed from DNA isolated from a pilot-plant biogas reactor operating at 55Â °C. The library was screened for starch-degrading enzymes, and one active clone was found. An open reading frame of 1,461Â bp encoding an [alpha]-amylase from an uncultured organism was identified. The amy13A gene was cloned in Escherichia coli, resulting in high-level expression of the recombinant amylase. The novel enzyme Amy13A showed the highest sequence identity (75Â %) to [alpha]-amylases from Petrotoga mobilis and Halothermothrix orenii. Amy13A is highly thermoactive, exhibiting optimal activity at 80Â °C, and it is also highly salt-tolerant, being active in 25Â % (w/v) NaCl. Amy13A is one of the few enzymes that tolerate high concentrations of salt and elevated temperatures, making it a potential candidate for starch processing under extreme conditions.[PUBLICATION ABSTRACT]