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Synthesis of α-galactooligosaccharides with α-galactosidase from Lactobacillus reuteri of canine origin



Crude cell-free extracts from Lactobacillus reuteri grown on cellobiose, maltose, lactose and raffinose were assayed for glycosidic activities. When raffinose was used as the carbon source, α-galactosidase was produced, showing the highest yield at the beginning of the stationary growth phase. A 64 kDa enzyme was purified by ultra- and gel filtration, and characterized for its hydrolytic and synthetic activity. Highest hydrolytic activity was found at pH 5.0 at 50 °C ( K M 0.55 mM, V max 0.80 μmol min −1 mg −1 of protein). The crude cell-free extract was further used in glycosyl transfer reactions to synthesize oligosaccharides from melibiose and raffinose. At a substrate concentration of 23% (w/v) oligosaccharide mixtures were formed with main products being a trisaccharide at 26% (w/w) yield from melibiose after 8 h and a tetrasaccharide at 18% (w/w) yield from raffinose after 7 h. Methylation analysis revealed the trisaccharide to be 6′ α-galactosyl melibiose and the tetrasaccharide to be stachyose. In both cases synthesis ceased when hydrolysis of the substrate reached 50%.



Applied Microbiology and BiotechnologySpringer Journals

Published: Dec 1, 2003

DOI: 10.1007/s00253-003-1426-0

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