Methods to Study the Structure and Catalytic Activity of cis-Splicing Inteins.
Methods to Study the Structure and Catalytic Activity of cis-Splicing Inteins.
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DOI：
10.1007/978-1-0716-0434-2_4
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发表时间：
2020
影响因子：
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通讯作者：
J. Zhao;Zhenming Du;Chunyu Wang;K. Mills
中科院分区：
文献类型：
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作者：
J. Zhao;Zhenming Du;Chunyu Wang;K. Mills
内含肽促进蛋白质剪接的自催化过程，内含肽会中断称为外显肽的侧翼多肽。蛋白质剪接的机制已通过大肠杆菌中的过表达进行了研究。大肠杆菌内含肽与非天然外显肽融合蛋白。内含肽可用于生成反应性 α-硫酯以及具有 N 末端 Cys 残基的蛋白质，以促进表达的蛋白质连接。因此，更详细地了解内含肽的功能可以对生物技术应用产生重大影响。在这里，我们提供生化方法来研究剪接活性和NMR方法来研究内含肽结构和催化机制。
The autocatalytic process of protein splicing is facilitated by an intein, which interrupts flanking polypeptides called exteins. The mechanism of protein splicing has been studied by overexpression inE. coliof intein fusion proteins with nonnative exteins. Inteins can be used to generate reactive α-thioesters, as well as proteins with N-terminal Cys residues, to facilitate expressed protein ligation. As such, a more detailed understanding of the function of inteins can have significant impact for biotechnology applications. Here, we provide biochemical methods to study splicing activity and NMR methods to study intein structure and the catalytic mechanism.