A thermoactive glucoamylase with biotechnological relevance from the thermoacidophilic Euryarchaeon Thermoplasma acidophilum

Abstract

A gene encoding an intracellular glucoamylase was identified in the genome of the extreme thermoacidophilic Archaeon Thermoplasma acidophilum. The gene taGA, consisting of 1,911 bp, was cloned and successfully expressed in Escherichia coli. The recombinant protein was purified 22-fold to homogeneity using heat treatment, anion-exchange chromatography, and gel filtration. Detailed analysis shows that the glucoamylase, with a molecular weight of 66 kDa per subunit, is a homodimer in its active state. Amylolytic activity was measured over a wide range of temperature (40-90 degrees C) and pH (pH 3.5-7) and was maximal at 75 degrees C and at acidic condition (pH 5). The recombinant archaeal glucoamylase uses a variety of polysaccharides as substrate, including glycogen and amylose. Maximal activity was measured towards amylopectin with a specific activity of 4.2 U/mg and increased almost threefold in the presence of manganese. Calcium ions have a pronounced effect on enzyme stability; in the presence of 5 mM CaCl(2), the half-life increased from 15 min to 2 h at 80 degrees C.

A Thermostable Glucoamylase from Bispora sp. MEY-1 with Stability over a Broad pH Range and Significant Starch Hydrolysis Capacity

“…Archaea growing at temperatures higher than 60°C is another promising microbial source of biocatalysts for industrial starch processing. Some glucoamylases from hyperthermophilic Sulfolobus [12], Thermoplasma [13], Picrophilus and Methanococcus [14] have been purified and characterized. But there are still many obstacles to be overcome, such as cultivation of hyperthermophilic archaea and poor heterogeneous expression.…”

Extracellular amylase production of a thermotolerant Fusarium sp: isolated from Eastern Nigerian soil

“…Other applications include use in the sewage treatment to reduce disposable solid content of the sludge and for pretreatment of animal feed to improve the digestibility (Kokab et al, 2003;Regulapati et al, 2007, Saxena et al, 2007. Starch is composed of two high molecular weight components, including amylose (15-25%), a linear polymer made up of α-1,4-linked glucopyranose residues and amylopectin (75-85%), a branched polymer made up of α-1,4 and α-1,6 linked glucopyranose residues (Dock et al, 2008). Starch degrading enzymes include endoamylases (α-amylase), exo-amylases (β-amylase), debranching enzymes and glycosyltransferases (Cereia et al, 2006, Khajeh et al, 2006.…”

Heterologous Expression and Characterization of an Alcohol Dehydrogenase from the Archeon Thermoplasma acidophilum

“…This feature could be explained by the fact that T. acidophilum and other thermoacidophilic organisms maintain low internal pHs [3,29] and as recently reported; a super-slow protein unfolding mechanism could be used as a central strategy to allow these proteins to function at extreme temperatures [30]. This has been observed for instance in native and heterologous proteins produced in T. acidophilum [31][32][33].…”