The ability of the beta-fructofuranosidase (EC 3.2.1.26) from Bifidobacterium lactis DSM 10140T to cleave a variety of fructooligosaccharides was characterised. We identified its gene on a cloned chromosomal DNA fragment by sequence similarity (69% identity) to the putative CscA protein encoded in the Bifidobacterium longum genome. The deduced amino acid sequence of 532 residues (59.4 kDa) appeared to be identical to the beta-fructofuranosidase from the same strain recently described by Ehrmann et al. (Curr. Microbiol. 2003, 46, 391-397). However, the characterisation of the heterologously expressed enzyme showed several discrepancies to the referred study. First, the B. lactis beta-fructofuranosidase gene was found to have 41% identity with CscA from E. coli in contrast to the 16% reported, therefore it was assigned to as CscA protein instead of BfrA. Second, we observed only low activity of the enzyme towards sucrose (6%) instead of the 100% previously reported. Instead, we measured highest activity (100%) of the enzyme with the oligofructose Raftilose as a substrate compared with the inulin of low degree of polymerisation Raftiline LS (29%) and the highly polymerised Raftiline HP (10%). Altogether, the enzyme showed high affinity to terminal beta(2-1) glycosyl linkages between fructose moieties. The Km values obtained for Raftilose, Raftiline LS and sucrose were 0.12, 7.08 and 8.37 mM, respectively, and V(max) values for the conversion to fructose were calculated to be 5, 21 and 17 micromol/min per mg of protein, respectively. Growth of B. lactis was supported by fructans of low degree of polymerisation (Raftilose and Raftiline LS), whereas we observed no growth with highly polymerised inulin (Raftiline HP).

中文翻译：

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乳酸双歧杆菌的重组β-果糖呋喃糖苷酶水解低聚果糖。

表征了来自乳双歧杆菌DSM 10140T的β-果糖呋喃糖苷酶（EC 3.2.1.26）切割多种低聚果糖的能力。我们通过与长双歧杆菌基因组中编码的假定的CscA蛋白序列相似性（69％一致性）在克隆的染色体DNA片段上鉴定了其基因。推导的532个残基（59.4 kDa）的氨基酸序列似乎与Ehrmann等人最近描述的同一菌株的β-果糖呋喃糖苷酶相同。（Curr.Microbiol.2003，46，391-397）。但是，异源表达的酶的表征与参考研究显示出一些差异。首先，发现乳酸双歧杆菌β-果糖呋喃糖苷酶基因与来自大肠杆菌的CscA具有41％的同一性，而据报道只有16％，因此，它被分配为CscA蛋白而不是BfrA。其次，我们观察到该酶对蔗糖的活性较低（6％），而不是先前报道的100％。相反，与低聚合度Raftiline LS（29％）和高聚合度Raftiline HP（10％）的菊粉相比，我们以低聚果糖Raftilose为底物测量了该酶的最高活性（100％）。总之，该酶对果糖部分之间的末端β（2-1）糖基键具有很高的亲和力。Raftilose，Raftiline LS和蔗糖获得的Km值分别为0.12、7.08和8.37 mM，计算得出的转化为果糖的V（max）值分别为每mg蛋白质5、21和17 micromol / min。 。B的生长