A thermostable alpha-galactosidase from Lactobacillus fermentum CRL722. Genetic characterization and main properties

CARRERA-SILVA, E.A.; SILVESTRONI, A.; LEBLANC, J.G.; PIARD, J.-C.; SAVOY DE GIORI, G.; SESMA, F.

CURRENT MICROBIOLOGY

Springer

Año: 2006 vol. 53 p. 374 - 378

0343-8651

The alpha-Galactosidase (alpha-Gal) enzyme, which is encoded by the melA gene, hydrolyzes alpha-1,6 galactoside linkages found in sugars such as raffinose and stachyose. These alpha-galacto-oligosaccharides (alpha-GOS), which are found in large quantities in vegetables such as soy, can cause gastrointestinal disorders in sensitive individuals since monogastric animals (including humans) do not posses alpha-Gal in the gut. The use of microbial alpha-Gal is a promising alternative to eliminate alpha-GOS in soy derived products. Using degenerate primers, the melA gene from Lactobacillus (L.) fermentum CRL 722 was identified. The complete genomic sequence of melA (2223 kbp) and of the genes flanking melA were obtained using a combination of techniques (conventional, uneven and reverse PCR, Southern hybridization) and shows strong similarities with the alpha-Gal gene of thermophilic microorganisms. The alpha-Gal from L. fermentum CRL 722 was purified from cell-free extracts using various techniques (ion exchange chromatography, salt precipitation, SDS-PAGE and ultra-filtration) and the main biochemical properties of the enzyme were determined. The enzyme was active at moderately high temperatures (55ºC), and stable at wide ranges of temperatures and pH. The thermostable alpha-Gal from L. fermentum CRL 722 could thus be used for technological applications such as the removal of alpha-GOS found in soy products. The complete melA gene could also be inserted in other microorganisms which can survive in the harsh conditions of the gut in order to degrade alpha-GOS in situ. Both strategies would improve the overall acceptability of soy-derived products by improving their nutritional value.