通过去除隐蔽翻译起始位点消除大肠杆菌中表达的截短重组蛋白。
Elimination of truncated recombinant protein expressed in Escherichia coli by removing cryptic translation initiation site.
作者信息
Jennings Matthew J, Barrios Adam F, Tan Song
机构信息
Center for Eukaryotic Gene Regulation, Department of Biochemistry and Molecular Biology, The Pennsylvania State University, University Park, PA 16802, USA.
Center for Eukaryotic Gene Regulation, Department of Biochemistry and Molecular Biology, The Pennsylvania State University, University Park, PA 16802, USA.
出版信息
Protein Expr Purif. 2016 May;121:17-21. doi: 10.1016/j.pep.2015.12.001. Epub 2015 Dec 29.
Undesirable truncated recombinant protein products pose a special expression and purification challenge because such products often share similar chromatographic properties as the desired full length protein. We describe here our observation of both full length and a truncated form of a yeast protein (Gcn5) expressed in Escherichia coli, and the reduction or elimination of the truncated form by mutating a cryptic Shine-Dalgarno or START codon within the Gcn5 coding region. Unsuccessful attempts to engineer in a cryptic translation initiation site into other recombinant proteins suggest that cryptic Shine-Dalgarno or START codon sequences are necessary but not sufficient for cryptic translation in E. coli.
不良的截短重组蛋白产物带来了特殊的表达和纯化挑战，因为这类产物通常与所需的全长蛋白具有相似的色谱特性。我们在此描述了在大肠杆菌中表达的酵母蛋白（Gcn5）的全长形式和截短形式的观察结果，以及通过突变Gcn5编码区内一个隐蔽的Shine-Dalgarno序列或起始密码子来减少或消除截短形式的情况。在其他重组蛋白中引入隐蔽翻译起始位点的尝试未成功，这表明隐蔽的Shine-Dalgarno序列或起始密码子序列对于大肠杆菌中的隐蔽翻译是必要的，但并不充分。
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