The effect of proline insertions on the thermostability of a barley α-glucosidase

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Abstract
The thermal stability of α-glucosidase is important because the conversion of starch to fermentable sugars during industrial production of ethanol (e.g. brewing, fuel ethanol production) typically takes place at temperatures of 65–73°C. In this study we investigate the thermostability of α-glucosidases from four plant species, compare their deduced amino acid sequences, and test the effect of substituting a proline for the residue present in the wild-type enzyme on the thermostability of α-glucosidase. The α-glucosidase from barley (Hordeum vulgare) was significantly less thermostable than the other three α-glucosidases. A comparison of the published deduced amino acid sequences of these four α-glucosidases revealed conserved proline residues in the three most thermostable α-glucosidases that were not found in the barley enzyme. Site-directed mutagenesis was done on recombinant barley α-glucosidase to create proteins with prolines at these conserved positions. The thermostability (T50) of one of these mutant enzymes, T340P, was 10°C higher than the non-mutated enzyme.

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