Intra- and extracellular β-galactosidases from Bifidobacterium bifidum and B-infantis:: Molecular cloning, heterologous expression, and comparative characterization
Intra- and extracellular β-galactosidases from Bifidobacterium bifidum and B-infantis:: Molecular cloning, heterologous expression, and comparative characterization
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DOI：
10.1128/aem.67.5.2276-2283.2001
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发表时间：
2001-05-01
影响因子：
4.4
通讯作者：
Stougaard, P
中科院分区：
文献类型：
--
作者：
Moller, PL;Jorgensen, F;Stougaard, P
从两歧双歧杆菌DSM 20215中分离并鉴定了3个β-半乳糖苷酶基因，从双歧杆菌DSM 20088中分离并鉴定了1个β-半乳糖苷酶基因。三个B。两歧属β-半乳糖苷酶彼此之间以及与先前公开的归类为家族2糖基水解酶的β-半乳糖苷酶之间显示出低程度的氨基酸序列相似性。同样，B。β-半乳糖苷酶与分类为家族42糖基水解酶的酶有较远的亲缘关系。来自B的一种酶。BIF 3 α-半乳糖苷酶的其它特殊特征是（i）活性酶的单体结构，包括1，752个氨基酸残基（188 kDa）和（ii）分子组织为N-末端β-半乳糖苷酶结构域和C-末端半乳糖结合结构域。另外两个B。双歧杆菌β-半乳糖苷酶和来自B的酶。α-淀粉酶是多聚体的细胞内酶，分子量分别与典型的家族2和家族42糖基水解酶相似。尽管大小、分子组成和氨基酸序列不同，但所有四种β-半乳糖苷酶都对β-D-半乳糖苷键的水解具有高度特异性，并且所有四种酶都能够以乳糖为底物进行半乳糖基转移。
Three beta -galactosidase genes from Bifidobacterium bifidum DSM20215 and one beta -galactosidase gene from Bifidobacterium infantis DSM20088 were isolated and characterized. The three B. bifidum beta -galactosidases exhibited a low degree of amino acid sequence similarity to each other and to previously published beta -galactosidases classified as family 2 glycosyl hydrolases. Likewise, the B. infantis beta -galactosidase was distantly related to enzymes classified as family 42 glycosyl hydrolases. One of the enzymes from B. bifidum, termed BIF3, is most probably an extracellular enzyme, since it contained a signal sequence which was cleaved off during heterologous expression of the enzyme in Escherichia coli, Other exceptional features of the BIF3 a-galactosidase were (i) the monomeric structure of the active enzyme, comprising 1,752 amino acid residues (188 kDa) and (ii) the molecular organization into an N-terminal beta -galactosidase domain and a C-terminal galactose binding domain. The other two B. bifidum beta -galactosidases and the enzyme from B. infantis were multimeric, intracellular enzymes with molecular masses similar to typical family 2 and family 42 glycosyl hydrolases, respectively. Despite the differences in size, molecular composition, and amino acid sequence, all four beta -galactosidases were highly specific for hydrolysis of beta -D-galatiosidic linkages, and all four enzymes were able to transgalactosylate with lactose as a substrate.