Department of Nutrition and Food Hygiene, West China School of Public Health, Sichuan University, Chengdu 610041, Sichuan, China.
Biomed Environ Sci. 2012 Apr;25(2):203-9. doi: 10.3967/0895-3988.2012.02.012.
This study is to examine the secretion effects of beta-galactosidase in Lactococcus lactis.
The usp45 and beta-galactosidase genes were cloned and inserted into plasmid pMG36e to obtain the recombinant plasmid pMG36e-usp-lacZ. This recombinant plasmid was transformed into both Escherichia coli DH5alpha and L. lactis MG1363. The enzyme activity, gene sequencing, SDS-PAGE and hereditary stability were assessed and studied.
The lacZ gene inserted into plasmids pMG36e-usp-lacZ was 99.37% similar to the GenBank sequence, and SDS-PAGE revealed an evident idio-strap at 116 KDa between L. lactis MG1363/pMG36e-usp-lacZ in both supernatant and cell samples. Beta-Galactosidase activity measured 0.225 U/mL in L. lactis pMG36e-usp-lacZ transformants, and its secretion rate was 10%. The plasmid pMG36e-usp-lacZ appeared more stable in MG1363.
The authors concluded that these new recombinant bacteria well expressed and secreted beta-galactosidase, indicating that the beta-galactosidase expression system was successfully constructed, and this might provide a new solution for management of lactose intolerance specifically and promote the use of gene-modified organisms as part of the food-grade plasmid in general.
本研究旨在检验β-半乳糖苷酶在乳球菌中的分泌效果。
克隆usp45 和β-半乳糖苷酶基因并插入质粒 pMG36e 中，得到重组质粒 pMG36e-usp-lacZ。将该重组质粒转化入大肠杆菌 DH5α 和乳球菌 MG1363 中。评估并研究酶活性、基因测序、SDS-PAGE 和遗传稳定性。
插入质粒 pMG36e-usp-lacZ 的 lacZ 基因与 GenBank 序列的相似度为 99.37%，SDS-PAGE 显示在乳球菌 MG1363/pMG36e-usp-lacZ 的上清液和细胞样品中，存在一条 116 kDa 的明显特征条带。L. lactis pMG36e-usp-lacZ 转化子中的β-半乳糖苷酶活性为 0.225 U/mL，其分泌率为 10%。质粒 pMG36e-usp-lacZ 在 MG1363 中更为稳定。
作者得出结论，这些新的重组菌能很好地表达和分泌β-半乳糖苷酶，表明β-半乳糖苷酶表达系统构建成功，这可能为乳糖不耐受的管理提供新的解决方案，同时促进将基因修饰生物体作为食品级质粒的一部分的应用。
