Total chemical synthesis of SUMO-2-Lys63-linked diubiquitin hybrid chains assisted by removable solubilizing tags.
Total chemical synthesis of SUMO-2-Lys63-linked diubiquitin hybrid chains assisted by removable solubilizing tags.
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DOI：
10.1039/c7sc00488e
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发表时间：
2017-05-01
期刊：
影响因子：
8.4
通讯作者：
Brik A
中科院分区：
文献类型：
--
作者：
Bondalapati S;Eid E;Mali SM;Wolberger C;Brik A
我们报告了四个不同的SUMO-2-Lys63连接的双泛素杂交链的第一个全化学合成，其中双泛素连接到SUMO中的不同赖氨酸。 小泛素样修饰蛋白（SUMO）是一种重要的细胞调控蛋白，参与细胞的转录和凋亡等重要的生物学过程。最近，发现含有SUMO-2连接到Lys63-二泛素的杂合SUMO-泛素链在DNA修复中起主要作用。尽管在了解这些混合链在DNA修复中的作用方面取得了一些进展，但仍有各种基本问题有待回答。为了进一步研究杂合SUMO-泛素链在DNA修复中的重要性，需要这些链的同质材料及其独特的类似物以可操作的数量。通过应用先进的蛋白质合成的化学策略，我们报告了四个不同的SUMO-2-Lys63连接的二泛素杂合链的第一次全化学合成，其中二泛素连接到SUMO中的不同赖氨酸。在这些合成中，证明了可去除的增溶标签的有用性，并且在可靠性和效率方面检查了两种不同的方法。在第一种方法中，聚精氨酸标签通过3，4-二氨基苯甲酸可切割接头连接到SUMO的C-末端，而在第二种方法中，我们通过苯乙酰氨基甲基接头连接标签，该接头可以被PdCl 2切割。这些不同策略之间的比较为这些类似物和其他蛋白质的未来放大制备提供了指导，这些蛋白质目前使用难以处理和纯化的合成肽中间体。SUMO-泛素杂合链的可用性为研究这些链在DNA修复和其他细胞过程中的作用开辟了新的机会。
We report the first total chemical synthesis of four different SUMO-2-Lys63-linked di-ubiquitin hybrid chains, in which the di-ubiquitin is linked to different lysines in SUMO. Small ubiquitin like modifier (SUMO) proteins are known to regulate many important cellular processes such as transcription and apoptosis. Recently, hybrid SUMO-ubiquitin chains containing SUMO-2 linked to Lys63-di-ubiquitin were found to play a major role in DNA repair. Despite some progress in understanding the role of these hybrid chains in DNA repair, there are various fundamental questions remaining to be answered. To further investigate the importance of hybrid SUMO-ubiquitin chains in DNA repair, the homogenous material of these chains, and their unique analogues, are needed in workable quantities. By applying advanced chemical strategies for protein synthesis, we report the first total chemical synthesis of four different SUMO-2-Lys63-linked di-ubiquitin hybrid chains, in which the di-ubiquitin is linked to different lysines in SUMO. In these syntheses, the usefulness of removable solubilizing tags is demonstrated, and two different approaches were examined in terms of reliability and efficiency. In the first approach, a poly-Arg tag was attached to the C-terminus of SUMO via a 3,4-diaminobenzoic acid cleavable linker, whereas in the second we attached the tag via a phenylacetamidomethyl linker, which can be cleaved by PdCl2. The comparison between these different strategies offers guidelines for future scale-up preparation of these analogues and other proteins, which currently use synthetic peptide intermediates that are difficult to handle and purify. The availability of the SUMO-ubiquitin hybrid chains opens up new opportunities for studying the role of these chains in DNA repair and other cellular processes.