Overproduction of beta-glucosidase in active form by an Escherichia coli system coexpressing the chaperonin GroEL/ES.
Overproduction of beta-glucosidase in active form by an Escherichia coli system coexpressing the chaperonin GroEL/ES.
复制标题
共表达伴侣蛋白 GroEL/ES 的大肠杆菌系统过量生产活性形式的 β-葡萄糖苷酶。
DOI：
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发表时间：
1998
影响因子：
2.1
通讯作者：
Yasushi Kawata
中科院分区：
文献类型：
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作者：
Sachiko Machida;Yong Yu;Satya P. Singh;Jong;Kiyoshi Hayashi;Yasushi Kawata
通过 GroEL/ES 的共表达，来自 Cellvibrio gilvus 的 β-葡萄糖苷酶在大肠杆菌中以可溶形式成功过量生产。如果没有GroEL/ES蛋白，大肠杆菌中过表达的β-葡萄糖苷酶占细胞总蛋白的大量(80%)，但局限于不溶性部分，在可溶性部分中检测到很少的活性。大肠杆菌 GroEL/ES 的共表达对 β-葡萄糖苷酶的正确折叠产生了巨大影响； 20% 的过表达酶以活性形式回收到可溶部分中。此外，GroEL/ES 和低诱导温度的协同效应导致总表达目标蛋白溶解 70%，活性提高 20 倍以上。 GroEL/ES 对来自根癌农杆菌的过表达 β-葡萄糖苷酶也观察到类似的效果。
beta-Glucosidase from Cellvibrio gilvus was successfully overproduced in soluble form in Escherichia coli, with the coexpression of GroEL/ES. Without the GroEL/ES protein, the beta-glucosidase overexpressed in E. coli constituted a huge amount (80%) of the total cellular protein, but was localized in the insoluble fraction, and little activity was detected in the soluble fraction. Coexpression of the E. coli GroEL/ES had a drastic impact on the proper folding of the beta-glucosidase; 20% of the overexpressed enzyme was recovered in the soluble fraction in active form. In addition, the synergistic effect of GroEL/ES and the low induction temperature led to 70% solubilization of the total expressed target protein and more than a 20-fold increase in activity. Similar effects of GroEL/ES were also observed on the overexpressed beta-glucosidase from Agrobacterium tumefaciens.