Production of a soluble and functional recombinant streptavidin in Escherichia coli
Production of a soluble and functional recombinant streptavidin in Escherichia coli
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DOI：
10.1006/prep.1998.0930
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发表时间：
1998-11-01
影响因子：
1.6
通讯作者：
Arosio, P
中科院分区：
文献类型：
--
作者：
Gallizia, A;de Lalla, C;Arosio, P
将链霉亲和素cDNA（15-159个残基）亚克隆到n端与t7标签（12个残基）融合的表达载体中。在大肠杆菌中发现了以可溶性、组装和活性形式表达蛋白质的条件。该蛋白通过两个简单的步骤纯化，即在75℃下加热和在亚胺生物素琼脂糖上亲和层析。纯化蛋白的产量为每升细菌培养70毫克。电子喷雾质谱分析表明，重组链霉亲和素具有预期的分子质量，无共价修饰。酶联免疫吸附试验和表面等离子体共振分析表明其功能与天然链霉亲和素相似。这似乎比报道的重组链亲和素生产方法有所改进，这些方法涉及蛋白质再生或使用真核表达系统。(C) 1998学术出版社。
The cDNA for streptavidin (residues 15-159) was subcloned into an expression vector in fusion at the N-terminus with the T7-tag (12 residues). Conditions were found to express the protein in Escherichia coli in a soluble, assembled, and active form. The protein was purified in two simple steps which involved heating at 75 degrees C and affinity chromatography on iminobiotin agarose. The purified protein was obtained in yields of 70 mg per liter of bacterial culture. Electron spray mass spectrometry analysis showed that the re combinant streptavidin had the expected molecular mass without covalent modifications. ELISA and surface plasmon resonance analyses showed it to be functionally analogous to the natural streptavidin. This appears to be an improvement over the reported methods of recombinant streptavidin production which involve protein renaturation or the use of eukaryotic expression systems. (C) 1998 Academic Press.