Quantitative analysis of in vitro ubiquitinated cyclin B1 reveals complex chain topology
Quantitative analysis of in vitro ubiquitinated cyclin B1 reveals complex chain topology
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DOI：
10.1038/ncb1436
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发表时间：
2006-07-01
影响因子：
21.3
通讯作者：
Gygi, Steven P.
中科院分区：
文献类型：
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作者：
Kirkpatrick, Donald S.;Hathaway, Nathaniel A.;Gygi, Steven P.
蛋白质泛素化调节许多细胞过程，包括蛋白质降解、信号转导、DNA修复和细胞分裂。在经典模型中，26S蛋白酶体的识别和降解需要一个通过Lys 48连接的均匀的多聚泛素链。在这里，我们使用了一个重建系统和定量质谱证明，细胞周期蛋白B1被复杂拓扑结构的泛素链修饰，而不是由同质的赖氨酸48连接的链。发现后期促进复合物将monoubiquitin连接到细胞周期蛋白B1上的多个赖氨酸残基上，然后通过泛素的多个赖氨酸残基（Lys 63，Lys 11和Lys 48）连接多聚泛素链延伸。这些异质性泛素链足以结合到泛素受体，以及降解的26 S蛋白酶体，即使当他们合成的突变体泛素缺乏赖氨酸48。总之，我们的观察扩展了什么可以被认为是一个足够的降解信号的背景，并提供独特的见解底物泛素化的机制。
Protein ubiquitination regulates many cellular processes, including protein degradation, signal transduction, DNA repair and cell division. In the classical model, a uniform polyubiquitin chain that is linked through Lys 48 is required for recognition and degradation by the 26S proteasome. Here, we used a reconstituted system and quantitative mass spectrometry to demonstrate that cyclin B1 is modified by ubiquitin chains of complex topology, rather than by homogeneous Lys 48-linked chains. The anaphase-promoting complex was found to attach monoubiquitin to multiple lysine residues on cyclin B1, followed by polyubiquitin chain extensions linked through multiple lysine residues of ubiquitin ( Lys 63, Lys 11 and Lys 48). These heterogeneous ubiquitin chains were sufficient for binding to ubiquitin receptors, as well as for degradation by the 26S proteasome, even when they were synthesized with mutant ubiquitin that lacked Lys 48. Together, our observations expand the context of what can be considered to be a sufficient degradation signal and provide unique insights into the mechanisms of substrate ubiquitination.