The Small Ubiquitin-like Modifier (SUMO) is a crucial post-translational modifier of proteins, playing a key role in various cellular functions. All SUMOs are synthesized as precursor proteins that must be proteolytically processed. However, the maturation process of cleaving the extending C-terminal tail, preceding SUMOylation of substrates, remains poorly understood, especially within cellular environments. Chemical protein synthesis coupled with cell delivery offers great opportunities to prepare SUMO analogues to investigate this process in vitro and in live cells. Applying this unique combination we show that SUMO2 analogues containing the native tail undergo rapid cleavage and nuclear localisation, while a Gly93Ala mutation impairs cleavage and alters localisation. Tail mutations (Val94Glu, Tyr95Ala) affected cleavage rates, highlighting roles in SUMO-SENP protease interactions. In cells, analogoues containing tail mutations underwent cleavage and subsequently incorporated into PML-NBs. These findings advance our understanding of SUMO2 maturation and provide a foundation for future studies of this process for different SUMO paralogues in various cell lines and tissues.

中文翻译：

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化学蛋白质合成与蛋白质细胞递送相结合揭示了 SUMO2 成熟过程的新见解


小泛素样修饰剂 （SUMO） 是蛋白质的重要翻译后修饰剂，在各种细胞功能中起关键作用。所有 SUMO 都是作为前体蛋白合成的，必须进行蛋白水解加工。然而，在底物 SUMO 化之前切割延伸的 C 末端尾部的成熟过程仍然知之甚少，尤其是在细胞环境中。化学蛋白质合成与细胞递送相结合，为制备 SUMO 类似物以在体外和活细胞中研究这一过程提供了绝佳的机会。应用这种独特的组合，我们表明包含天然尾巴的 SUMO2 类似物经历快速切割和核定位，而 Gly93Ala 突变损害切割并改变定位。尾部突变 （Val94Glu、Tyr95Ala） 影响切割率，突出了在 SUMO-SENP 蛋白酶相互作用中的作用。在细胞中，含有尾部突变的类似物发生切割，随后掺入 PML-NBs 中。这些发现促进了我们对 SUMO2 成熟的理解，并为未来研究各种细胞系和组织中不同 SUMO 旁系同源物的这一过程奠定了基础。