The cloning, purification and characterisation of a cold-active β-galactosidase from the psychrotolerant Antarctic bacterium Arthrobacter sp C2-2
The cloning, purification and characterisation of a cold-active β-galactosidase from the psychrotolerant Antarctic bacterium Arthrobacter sp C2-2
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DOI：
10.1016/s0141-0229(03)00211-4
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发表时间：
2003-11-05
影响因子：
3.4
通讯作者：
Russell, NJ
中科院分区：
文献类型：
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作者：
Karasová-Lipovová, P;Strnad, H;Russell, NJ
革兰氏阳性南极细菌节杆菌C2-2含有两种，可能是三种β-半乳糖苷酶的低温活性同工酶。对C2-2-1同工酶进行了克隆、纯化和表征。该β-半乳糖苷酶被分类为糖苷酶家族2的成员。它是一种同源四聚体酶，每个亚基由1023个氨基酸组成，并且对作为底物的乳糖显示出很大的活性。C2-2-1同工酶与其他β-半乳糖苷酶（包括来自一些密切相关的细菌的酶）相比，具有特别的低温活性，在10 ℃时保持20%的活性（与最大值相比）。以乳糖为底物，纯化酶的最适反应温度为40 ℃。该酶特别不耐热，在50 ℃下10分钟内失去所有活性。该酶的等电点为5.9。二硫苏糖醇和Mg ~（2+）是强激活剂，而Cu ~（2+）、Al ~（3+）和Tris是强抑制剂。该酶表现出转糖基化能力，在15 ℃下10 h后形成最高浓度的三聚氰胺（34 mM），这与商业酶的活性相当。因此，节杆菌属C2-2的C2-2-1，1，β-半乳糖苷酶同工酶具有特别的优点，即它可以用作在冷藏温度下生产乳糖减少的乳制品的生物技术工具。(C)2003年爱思唯尔公司All rights reserved.
The Gram-positive Antarctic bacterium Arthrobacter sp. C2-2 contains two, possibly three cold-active isoenzymes of beta-galactosidase. The C2-2-1 isoenzyme was cloned, purified and characterised. This beta-galactosidase was classified as being a member of the Family 2 of glycosidases. It is a homotetrameric enzyme, each subunit being composed of 1023 arnino acids and it shows great activity towards lactose as a substrate. The C2-2-1 isoenzyme is particularly cold-active, compared to other beta-galactosidases including those from some closely-related bacteria, retaining 20% of activity at 10degreesC compared with maximum values. The temperature optimum of the purified enzyme was 40degreesC using lactose as the substrate. The enzyme is particularly thermolabile, losing all activity within 10 min at 50degreesC. The isoelectric point of the enzyme was 5.9. Dithiothreitol and Mg2+ ions were strong activators, whereas Cu2+, Al3+ and Tris were strong inhibitors of activity. The enzyme exhibited transglycosylation ability and the highest concentration of trisaccharides (34 mM) was formed after 10h at 15degreesC, which is an activity comparable with that of the commercial enzymes. Therefore, the C2-2-1,1,beta-galactosidase isoenzyme of Arthrobacter sp. C2-2 has the particular advantage that it could be used as a biotechnological tool in the production of lactose-reduced dairy products at refrigeration temperatures. (C) 2003 Elsevier Inc. All rights reserved.