A highly thermoactive and salt-tolerant α-amylase isolated from a pilot-plant biogas reactor

Appl Microbiol Biotechnol. 2013 Apr;97(7):2971-8. doi: 10.1007/s00253-012-4194-x. Epub 2012 Jun 29.

Abstract

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 55 °C. The library was screened for starch-degrading enzymes, and one active clone was found. An open reading frame of 1,461 bp encoding an α-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 (75%) to α-amylases from Petrotoga mobilis and Halothermothrix orenii. Amy13A is highly thermoactive, exhibiting optimal activity at 80 °C, and it is also highly salt-tolerant, being active in 25% (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.

MeSH terms

  • Amino Acid Sequence
  • Biofuels
  • Bioreactors*
  • Cloning, Molecular
  • Enzyme Stability
  • Escherichia coli / genetics
  • Gene Expression
  • Metagenome*
  • Models, Molecular
  • Molecular Sequence Data
  • Protein Conformation
  • Recombinant Proteins / chemistry
  • Recombinant Proteins / genetics
  • Recombinant Proteins / metabolism
  • Sequence Homology, Amino Acid
  • Sodium Chloride / metabolism
  • Starch / metabolism
  • Temperature
  • alpha-Amylases / chemistry
  • alpha-Amylases / genetics*
  • alpha-Amylases / metabolism*

Substances

  • Biofuels
  • Recombinant Proteins
  • Sodium Chloride
  • Starch
  • alpha-Amylases