Chemical Genomics Centre, Max Planck Institute of Molecular Physiology, Dortmund, Germany.
Department of Chemistry and Chemical Biology, TU Dortmund University, Dortmund, Germany.
Nat Chem Biol. 2023 Nov;19(11):1394-1405. doi: 10.1038/s41589-023-01388-1. Epub 2023 Jul 13.
Ubiquitin and ubiquitin-like proteins typically use distinct machineries to facilitate diverse functions. The immunosuppressive ubiquitin-like protein Fubi is synthesized as an N-terminal fusion to a ribosomal protein (Fubi-S30). Its proteolytic maturation by the nucleolar deubiquitinase USP36 is strictly required for translationally competent ribosomes. What endows USP36 with this activity, how Fubi is recognized and whether other Fubi proteases exist are unclear. Here, we report a chemical tool kit that facilitated the discovery of dual ubiquitin/Fubi cleavage activity in USP16 in addition to USP36 by chemoproteomics. Crystal structures of USP36 complexed with Fubi and ubiquitin uncover its substrate recognition mechanism and explain how other deubiquitinases are restricted from Fubi. Furthermore, we introduce Fubi C-terminal hydrolase measurements and reveal a synergistic role of USP16 in Fubi-S30 maturation. Our data highlight how ubiquitin/Fubi specificity is achieved in a subset of human deubiquitinases and open the door to a systematic investigation of the Fubi system.
泛素和泛素样蛋白通常使用不同的机制来促进不同的功能。免疫抑制性泛素样蛋白 Fubi 作为核糖体蛋白（Fubi-S30）的 N 端融合物合成。其核仁去泛素酶 USP36 的蛋白水解成熟对于翻译功能完整的核糖体是严格必需的。USP36 具有这种活性的原因是什么，Fubi 如何被识别，以及是否存在其他 Fubi 蛋白酶尚不清楚。在这里，我们报告了一个化学工具包，通过化学蛋白质组学发现了 USP16 除了 USP36 之外还具有双重泛素/Fubi 切割活性。USP36 与 Fubi 和泛素复合物的晶体结构揭示了其底物识别机制，并解释了其他去泛素酶如何受到限制而不能作用于 Fubi。此外，我们引入了 Fubi C 端水解酶测量，并揭示了 USP16 在 Fubi-S30 成熟过程中的协同作用。我们的数据突出了在人类去泛素酶的亚类中如何实现泛素/Fubi 的特异性，并为 Fubi 系统的系统研究打开了大门。
