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


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Abstract

AimsTo characterize the β-fructofuranosidase of Bifidobacterium infantis ATCC 15697 and to compare it with other bacterial β-fructofuranosidases.Methods and ResultsThe β-fructofuranosidase of B. infantis ATCC 15697 was purified 46·8 times over the crude extract by anion exchange chromatography, ultrafiltration and gel filtration. The sequence of 15 amino acid residues of the NH2 terminal was determined. This enzyme was a monomeric protein (Mr 70 kDa) with β-fructofuranosidase and invertase activities. The isoelectric point was pH 4·3, the optimum pH 6·0 and pKas (4·5 and 7·2) of two active groups were obtained. The activities were inhibited by Hg2+ and p-chloromercuribenzoic acid (pCMB). The optimal temperature was 37 °C and activities were unstable at 55 °C. β-fructofuranosidase activity was more efficient than that of invertase with Vm/Km ratios of 0·65 and 0·025 × 10−3 l min−1 mg−1, respectively. The enzyme catalyses the hydrolysis of fructo-oligosaccharides, sucrose and inulin at relative velocities of 100, 10 and 6, respectively.ConclusionsThe enzyme of B. infantis ATCC 15697 is an exo-inulinase which has β-fructofuranosidase and invertase activities. This protein was different from the β-fructofuranosidase of another strain of B. infantis (B. infantis JCM no. 7007).Significance and Impact of the StudyA better knowledge of bacterial β-fructofuranosidases, especially from bifidobacteria, has been gained.

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