Recent advances in GFP folding reporter and split-GFP solubility reporter technologies.: Application to improving the folding and solubility of recalcitrant proteins from Mycobacterium tuberculosis
Recent advances in GFP folding reporter and split-GFP solubility reporter technologies.: Application to improving the folding and solubility of recalcitrant proteins from Mycobacterium tuberculosis
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DOI：
10.1007/s10969-005-5247-5
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发表时间：
2005-09-01
期刊：
影响因子：
--
通讯作者：
Waldo, Geoffrey S.
中科院分区：
文献类型：
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作者：
Cabantous, Stephanie;Pedelacq, Jean-Denis;Waldo, Geoffrey S.
我们已经改进了我们的绿色荧光蛋白（GFP）折叠报告基因技术[Waldo et al.，等人（1999）Nat. Biotechnol. 17，691-695]以从结核分枝杆菌进化出柠檬酸盐蛋白。将靶蛋白插入GFP的支架中，消除了由靶RNA中内部隐蔽核糖体结合位点的截短蛋白变体的表达引起的假阳性假象。同时，我们开发了一种新的基于GFP微结构域的荧光蛋白定量标记和检测系统。这种在体内和体外都起作用的分裂-GFP系统适用于蛋白质表达和溶解度的高通量测定[Cabantous等人，等人（2005）Nat. Biotechnol. 23，102-107]。GFP折叠报告基因和split-GFP技术共同提供了一个用于操纵和改善蛋白质折叠和溶解度的综合系统。
We have improved our green fluorescent protein (GFP) folding reporter technology [Waldo et al., (1999) Nat. Biotechnol. 17, 691-695] to evolve recalcitrant proteins from Mycobacterium tuberculosis. The target protein is inserted into the scaffolding of the GFP, eliminating false-positive artifacts caused by expression of truncated protein variants from internal cryptic ribosome binding sites in the target RNA. In parallel, we have developed a new quantitative fluorescent protein tagging and detection system based on micro-domains of GFP. This split-GFP system, which works both in vivo and in vitro, is amenable to high-throughput assays of protein expression and solubility [Cabantous et al., (2005) Nat. Biotechnol. 23, 102-107]. Together, the GFP folding reporter and split-GFP technologies offer a comprehensive system for manipulating and improving protein folding and solubility.