PURIFICATION AND CHARACTERIZATION OF NOVEL GLYCOSIDASES FROM THE BACTERIAL GENUS XANTHOMONAS
PURIFICATION AND CHARACTERIZATION OF NOVEL GLYCOSIDASES FROM THE BACTERIAL GENUS XANTHOMONAS
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DOI：
10.1093/glycob/5.1.19
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发表时间：
1995-02-01
期刊：
影响因子：
4.3
通讯作者：
LANDRY, D
中科院分区：
文献类型：
--
作者：
WONGMADDEN, ST;LANDRY, D
利用外切糖苷酶对低聚糖进行酶分析已成为确定糖链序列和结构的有力工具。这些方法的主要限制是缺乏高纯度和良好特性的酶。利用荧光标记的碳水化合物底物和TLC，我们开发了一种鉴定具有新特异性的糖苷酶的方法，这种筛选方法导致发现黄单胞菌属细菌是外糖苷酶的丰富来源。从Xanthomonas manihotis中分离和鉴定了8种新的外糖苷酶，并纯化了一种新的β-N-乙酰氨基葡萄糖苷酶，与前面描述的酶不同，它可以在不切割N-乙酰氨基半乳糖残基的情况下裂解N-乙酰氨基葡萄糖。一个新的β-半乳糖苷酶被分离出来，它优先水解β(1->；3)半乳糖基。三个α-甘露糖苷酶已经被分离出来，它们在高甘露糖寡糖结构的分析中是有用的试剂：α1-3，6甘露糖苷酶，α1-6甘露糖苷酶和α1-2，3甘露糖苷酶。纯化的α1-3，6半乳糖苷酶不能降解末端的α1-4半乳糖残基，两种岩藻糖苷酶，即α1-3，4岩藻糖苷酶和α1-2岩藻糖苷酶与其他来源的酶相似，它们共同为测定复杂碳水化合物的序列提供了强有力的试剂。同样重要的是，它们在选择性地去除特定糖类残留物，从而产生用于分析寡糖的生物作用的新型碳水化合物方面也是有用的。
Enzymatic analysis of oligosaccharides using exoglycosidases has become a powerful tool for determining the sequence and structure of sugar chains. The principal limitation to these methods has been the lack of highly purified and wellcharacterized enzymes. Using fluorescently labelled carbohydrate substrates and TLC, we have developed a method to identify glycosidases with novel specificities, This screening method led to the discovery that bacteria of the genus Xanthomonas are a rich source of exoglycosidases. From Xanthomonas manihotis, eight novel exoglycosidases have been isolated and characterized, A novel beta-N-acetylglucosaminidase has been purified that, unlike those previously described, will cleave N-acetylglucosamine without cleaving N-acetylgalactosamine residues. A novel beta-galactosidase has been isolated that preferentially hydrolyses beta(1-->3) galactosyl linkages, Three alpha-mannosidases have been isolated that serve as useful reagents in the analysis of high-mannose oligosaccharide structures: alpha 1-3,6 mannosidase, alpha 1-6 mannosidase and alpha 1-2,3 mannosidase. An alpha 1-3,6 galactosidase has been purified that does not hydrolyse terminal alpha 1-4 galactose residues, Two fucosidases, alpha 1-3,4 fucosidase and alpha 1-2 fucosidase, are similar to enzymes purified from other sources, Together, these glycosidases provide powerful reagents for determining the sequence of complex carbohydrates. Equally important is their usefulness in selectively removing specific sugar residues and thereby creating novel carbohydrates for analysing the biological roles of oligosaccharides.