Bonacci Thomas, Bolhuis Derek L, Brown Nicholas G, Emanuele Michael J
Department of Pharmacology and Lineberger Comprehensive Cancer Center, The University of North Carolina at Chapel Hill, Chapel Hill, North Carolina, USA.
Department of Biochemistry and Biophysics and Lineberger Comprehensive Cancer Center, The University of North Carolina at Chapel Hill, Chapel Hill, North Carolina, USA.
J Biol Chem. 2025 May 26;301(7):110288. doi: 10.1016/j.jbc.2025.110288.
The ubiquitin-like protein ISG15 is activated in response to type 1 interferons, and its conjugation to proteins regulates the response to bacterial and viral infection. Its subsequent deconjugation, which is broadly achieved by the human enzyme USP18, critically controls interferon signaling and the defense against pathogens. However, the molecular determinants underlying USP18 specificity for ISG15 remain elusive. To identify such features, we took advantage of USP18's paralog USP41, which has a strikingly similar catalytic domain and yet lacks deISGylating activity. By performing a comparative sequence analysis coupled with biochemical and enzymatic assays, we identified hallmarks specific to USP18 that are critical for its enzymatic function and ISG15 recognition. Accordingly, AlphaFold-guided analysis suggests that these features mediate USP18-ISG15 interactions, underlining their importance for deISGylating activities. Thus, our results reveal important mechanistic insights into USP18-mediated ISG15 hydrolysis and could inform the development of deISGylase inhibitors relevant to infection and other interferon-related diseases.
类泛素蛋白ISG15在1型干扰素的作用下被激活，其与蛋白质的缀合作用调节对细菌和病毒感染的反应。随后它的去缀合作用（主要由人类酶USP18实现）严格控制干扰素信号传导以及对病原体的防御。然而，USP18对ISG15特异性的分子决定因素仍然不清楚。为了确定这些特征，我们利用了USP18的旁系同源物USP41，它具有惊人相似的催化结构域，但缺乏去ISG化活性。通过进行比较序列分析并结合生化和酶学分析，我们确定了USP18特有的特征，这些特征对其酶功能和ISG15识别至关重要。因此，基于AlphaFold的分析表明，这些特征介导了USP18与ISG15的相互作用，突出了它们对去ISG化活性的重要性。因此，我们的结果揭示了关于USP18介导的ISG15水解的重要机制见解，并可能为与感染和其他干扰素相关疾病相关的去ISGylase抑制剂的开发提供信息。
