Characterization of a purified beta-fructofuranosidase from Bifidobacterium infantis ATCC 15697

Abstract

Aims: To characterize the b-fructofuranosidase of Bifidobacterium infantis ATCC 15697 and to compare it with other bacterial b-fructofuranosidases. Methods and Results: The b-fructofuranosidase of B. infantis ATCC 15697 was purified 46AE8 times over the crude extract by anion exchange chromatography, ultrafiltration and gel filtration. The sequence of 15 amino acid residues of the NH 2 terminal was determined. This enzyme was a monomeric protein (M r 70 kDa) with b-fructofuranosidase and invertase activities. The isoelectric point was pH 4AE3, the optimum pH 6AE0 and pKas (4AE5 and 7AE2) of two active groups were obtained. The activities were inhibited by Hg 2+ and p-chloromercuribenzoic acid (pCMB). The optimal temperature was 37°C and activities were unstable at 55°C. b-fructofuranosidase activity was more efficient than that of invertase with V m ⁄ K m ratios of 0AE65 and 0AE025 · 10 )3 l min )1 mg )1 , respectively. The enzyme catalyses the hydrolysis of fructooligosaccharides, sucrose and inulin at relative velocities of 100, 10 and 6, respectively. Conclusions: The enzyme of B. infantis ATCC 15697 is an exo-inulinase which has b-fructofuranosidase and invertase activities. This protein was different from the b-fructofuranosidase of another strain of B. infantis (B. infantis JCM no. 7007). Significance and Impact of the Study: A better knowledge of bacterial b-fructofuranosidases, especially from bifidobacteria, has been gained.

Biochemical properties of an extracellular β-D-fructofuranosidase II produced by Aspergillus phoenicis under Solid-Sate Fermentation using soy bran as substrate

“…This temperature optimum was higher than that reported for the enzyme from Bifidobacterium infantis ATCC 15697 (Warchol et al 2002), Lactobacillus reuteri (Ginés et al 2000), Fusarium solani (Bhatti et al 2006), Aspergillus niger IMI303386 (Nguyen et al 2005) and A. oryzae KB (Kurakake et al 2008), and similar with that of the β-fructofuranosidases from Aspergillus niger ATCC 20611 and Aureobasidium sp. ATCC 20524 (Yoshikawa et al 2007).…”

Bifidobacterium longum Requires a Fructokinase (Frk; ATP: d -Fructose 6-Phosphotransferase, EC 2.7.1.4) for Fructose Catabolism

“…Bioinformatic analyses complement biochemical data reporting the purification and characterization of Bifidobacterium sp. enzymes that are capable of hydrolyzing various glycosidic bonds from plant-derived carbohydrates and are often inducible by the released monomeric molecules (15,22,29,33,41,48,56,58,63). Moreover, several studies conducted on fructose-containing polymers as potential selective substrates for colonic bacteria have provided evidence that bifidobacteria are able to ferment these carbohydrates, in particular the short linear chains of ␤-2-1-linked fructosyl units (7,24,30,36,37).…”

Carbohydrate metabolism in Bifidobacteria

“…lactis [26,48], B. breve UCC2003 [113] and B. longum subsp. infantis [47,157]. Most, but not all, of the characterized b-fructofuranosidases exhibit hydrolytic activity towards various fructooligosaccharides, sucrose and inulin.…”