Discovery and characterization of non-canonical E2 conjugating enzymes
Discovery and characterization of non-canonical E2 conjugating enzymes
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DOI：
10.1101/2023.03.05.531151
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发表时间：
2023-03
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影响因子：
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通讯作者：
S. Rehman;Elena Di Nisio;Chiara Cazzaniga;O. Antico;A. Knebel;Clare Johnson;Frederic Lamoliatte;R. Negri;Miratul Muqit MK;V. D. Cesare
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作者：
S. Rehman;Elena Di Nisio;Chiara Cazzaniga;O. Antico;A. Knebel;Clare Johnson;Frederic Lamoliatte;R. Negri;Miratul Muqit MK;V. D. Cesare
E2结合酶（E2）在导致泛素与底物连接的酶级联反应中发挥核心作用。这一过程被称为泛素化，是维持细胞稳态的基础，几乎影响所有细胞过程。通过与多个E3连接酶相互作用，E2指导细胞内的泛素化景观。自发现以来，泛素化一直被认为是一种特异性靶向赖氨酸侧链的翻译后修饰（典型泛素化）。我们使用MALDI-TOF质谱法来发现和表征E2家族，其相反能够将泛素缀合至丝氨酸和/或苏氨酸。我们采用蛋白质建模和预测工具来确定这些E2用于与泛素及其底物相互作用的催化决定簇。我们的研究结果加入了最近的文献流，挑战泛素化的定义作为一个精致的赖氨酸特异性修饰，并提供了重要的见解缺失的E2元件负责非典型的泛素化。挑逗E2结合酶（E2）在泛素与其底物的连接中起着重要作用。大多数E2可以在泛素C-末端和底物上存在的赖氨酸之间形成异肽键。我们鉴定了E2家族，UBE 2 Q1和UBE 2 Q2，能够靶向赖氨酸以外的氨基酸。目前还不知道它们的作用机制和它们靶向的底物，尽管UBE 2 Q1的基因切除在小鼠中产生了大量的不育。在这里，我们回答的问题是什么关键残留物下的特殊活动。我们发现UBE 2 Q1靶向高尔基体驻留蛋白β-1，4-半乳糖基转移酶1的无赖氨酸胞质结构域，为真核细胞中非典型泛素化的作用提供了一个有趣的先例。
E2 conjugating enzymes (E2s) play a central role in the enzymatic cascade that leads to the attachment of ubiquitin to a substrate. This process, termed ubiquitylation is fundamental for maintaining cellular homeostasis and impacts almost all cellular process. By interacting with multiple E3 ligases, E2s direct the ubiquitylation landscape within the cell. Since its discovery, ubiquitylation has been regarded as a post-translational modification that specifically targets lysine side chains (canonical ubiquitylation). We used MALDI-TOF Mass Spectrometry to discover and characterize a family of E2s that are instead able to conjugate ubiquitin to serine and/or threonine. We employed protein modelling and prediction tools to identify the catalytic determinants that these E2s use to interact with ubiquitin as well as their substrates. Our results join a stream of recent literature that challenges the definition of ubiquitylation as an exquisitely lysine-specific modification and provide crucial insights into the missing E2 element responsible for non-canonical ubiquitylation. Teaser E2 conjugating enzymes (E2s) play a fundamental role in the attachment of ubiquitin to its substrate. Most E2s can form an isopeptide bond between the ubiquitin C- terminus and a lysine present on the substrate. We identified a family of E2s, UBE2Q1 and UBE2Q2, able to target amino acids other than lysine. Currently nothing is known about their mechanism of action and what substrates they are targeting, even though genetic ablation of UBE2Q1 produce substantial infertility in mice. Here we answer the question about what the key residues beneath their peculiar activity are. We discovered that UBE2Q1 target the lysine-free cytoplasmic domain of the Golgi resident protein Beta-1,4-galactosyltransferase 1, providing an interesting precedent for the role of non-canonical ubiquitylation in eukaryotic cells.