蛋白质-寡核苷酸缀合物作为 DNA-蛋白质交联修复蛋白酶的模型底物。
Protein-oligonucleotide conjugates as model substrates for DNA-protein crosslink repair proteases.
机构信息
Department of Biochemistry, Ludwig Maximilians University, 81377 Munich, Germany.
Gene Center, Ludwig Maximilians University, 81377 Munich, Germany.
出版信息
STAR Protoc. 2021 Jun 15;2(2):100591. doi: 10.1016/j.xpro.2021.100591. eCollection 2021 Jun 18.
Covalent DNA-protein crosslinks (DPCs) have emerged as pervasive sources of genome instability. DPCs are targeted for repair by DNA-dependent proteases of the Wss1/SPRTN family. However, understanding how these enzymes achieve specificity has been hampered by the lack of suitable model substrates. Here, we describe the generation of defined protein-oligonucleotide conjugates as DPC model substrates, which enable the analysis of DPC proteases in activity and binding assays. For complete details on the use and execution of this protocol, please refer to Reinking et al. (2020).
共价 DNA-蛋白质交联物 (DPCs) 已成为基因组不稳定性的普遍来源。DPCs 是 Wss1/SPRTN 家族的 DNA 依赖性蛋白酶的修复靶标。然而，由于缺乏合适的模型底物，这些酶如何实现特异性一直受到阻碍。在这里，我们描述了作为 DPC 模型底物的定义蛋白 - 寡核苷酸缀合物的生成，这使得可以在活性和结合测定中分析 DPC 蛋白酶。有关此方案的使用和执行的完整详细信息，请参阅 Reinking 等人。(2020)。
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