Identification, cloning and expression of a cold-active -galactosidase from a novel Arctic bacterium, Alkalilactibacillus ikkense

被引:33
作者
Schmidt, Mariane [1 ]
Stougaard, Peter [1 ]
机构
[1] Univ Copenhagen, Fac Life Sci, Dept Agr & Ecol, DK-1168 Copenhagen, Denmark
关键词
Alkalilactibacillus; -galactosidase; cold-active; psychrophilic; lactase; BETA-GALACTOSIDASE; STRAIN; PURIFICATION; ENZYMES; LACTOSE;
D O I
10.1080/09593331003677872
中图分类号
X [环境科学、安全科学];
学科分类号
08 ; 0830 ;
摘要
A novel, cold-active -galactosidase was isolated from an Arctic Gram-positive bacterium, Alkalilactibacillus ikkense. The corresponding gene was cloned and expressed as an active enzyme in Escherichia coli. Denaturing gel electrophoresis of both the native and the recombinant -galactosidase showed a monomeric molecular weight of 115-120 kDa. Analysis of the DNA sequence showed sequence similarity to known Glycosyl Hydrolase Family 2 -galactosidases from the genera Bacillus, Paenibacillus, Geobacillus, and Lactobacillus. The -galactosidase from this study was purified and shown to be highly active at low temperatures with more than 60% of its maximal activity maintained at 0 degrees C. The apparent optimal activity was observed at temperatures between 20 degrees C and 30 degrees C and at pH 8. The purified recombinant enzyme was stable without stabilizing agents for more than 100 hours at temperatures at and below 10 degrees C. At temperatures 40 degrees C and above, the -galactosidase was irreversibly inactivated within 10 minutes. When lactose was present in substantial amounts, the enzyme displayed transgalactosylation activity. Comparison of the -galactosidase with a commercially available enzyme showed that the conversion rate of the A. ikkense enzyme was approximately two-fold higher at temperatures between 0 degrees C and 20 degrees C.
引用
收藏
页码:1107 / 1114
页数:8
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