Modification of aniline containing proteins using an oxidative coupling strategy
Modification of aniline containing proteins using an oxidative coupling strategy
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DOI：
10.1021/ja064088d
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发表时间：
2006-12-13
影响因子：
15
通讯作者：
Francis, Matthew B.
中科院分区：
文献类型：
--
作者：
Hooker, Jacob M.;Esser-Kahn, Aaron P.;Francis, Matthew B.
基于氧化偶联途径对苯胺类化合物的化学选择性修饰，发展了一种新的生物偶联反应。通过赖氨酸酰化反应或通过使用天然化学连接技术将芳基胺安装在蛋白质底物的表面上。在NaIO_4缓冲溶液中，苯胺与N，N-二烷基-N’-酰基-对苯二胺的芳环迅速偶联。使用ESI-MS并通过与小分子类似物的比较来确认反应产物的身份。对照实验表明，没有天然氨基酸参与反应。发现所得生物缀合物在pH 4至pH 11的水解和许多常用的氧化剂、还原剂和亲核试剂的存在下是稳定的。合成了一种苯二胺荧光试剂，并在绿色荧光蛋白的C-末端添加了一条PEG链，用于苯胺标记蛋白的选择性检测。当与用于将非天然氨基酸掺入蛋白质中的技术相结合时，这种生物正交偶联方法应该证明对于需要高度标记特异性的许多应用是有用的。
A new bioconjugation reaction has been developed based on the chemoselective modification of anilines through an oxidative coupling pathway. Aryl amines were installed on the surface of protein substrates through lysine acylation reactions or through the use of native chemical ligation techniques. Upon exposure to NaIO4in aqueous buffer, the anilines coupled rapidly to the aromatic rings ofN,N-dialkyl-N‘-acyl-p-phenylenediamines. The identities of the reaction products were confirmed using ESI-MS and through comparison to small molecule analogs. Control experiments indicated that none of the native amino acids participated in the reaction. The resulting bioconjugates were found to be stable toward hydrolysis from pH 4 to pH 11 and in the presence of many commonly used oxidants, reductants, and nucleophiles. A fluorescent phenylenediamine reagent was synthesized for the selective detection of aniline labeled proteins in mixtures, and the reaction was used to append the C-terminus of the green fluorescent protein with a single PEG chain. When combined with techniques for the incorporation of unnatural amino acids into proteins, this bioorthogonal coupling method should prove useful for a number of applications requiring a high degree of labeling specificity.