Department of Chemistry, College of Arts and Science, University at Albany, State University of New York, 1400 Washington Avenue, Albany, NY-12222, USA.
The RNA Institute, College of Arts and Science, University at Albany, State University of New York, 1400 Washington Avenue, Albany, NY-12222, USA.
Chembiochem. 2022 Aug 3;23(15):e202200089. doi: 10.1002/cbic.202200089. Epub 2022 Jun 21.
Natural RNA modifications diversify the structures and functions of existing nucleic acid building blocks. Geranyl is one of the most hydrophobic groups recently identified in bacterial tRNAs. Selenouridine synthase (SelU, also called mnmH) is an enzyme with a dual activity which catalyzes selenation and geranylation in tRNAs containing 2-thiouridine using selenophosphate or geranyl-pyrophosphate as cofactors. In this study, we explored the in vitro geranylation process of tRNA anticodon stem loops (ASL) mediated by SelU and showed that the geranylation activity was abolished when U was mutated to A (ASL-tRNA to ASL-tRNA ). By examining the SelU cofactor geranyl-pyrophosphate (gePP) and its analogues, we found that only the geranyl group, but not dimethylallyl- and farnesyl-pyrophosphate with either shorter or longer terpene chains, could be incorporated into ASL. The degree of tRNA geranylation in the end-point analysis for SelU follows the order of ASL ASL >ASL . These findings suggest a putative mechanism for substrate discrimination by SelU and reveal key factors that might influence its enzymatic activity. Given that SelU plays an important role in bacterial translation systems, inhibiting this enzyme and targeting its geranylation and selenation pathways could be exploited as a promising strategy to develop SelU-based antibiotics.
天然 RNA 修饰使现有的核酸构建块的结构和功能多样化。香叶基是最近在细菌 tRNA 中鉴定出的最疏水基团之一。硒代尿嘧啶合酶（SelU，也称为 mnmH）是一种具有双重活性的酶，它可以使用硒代磷酸或香叶基焦磷酸作为辅助因子，催化含有 2-硫代尿嘧啶的 tRNA 的硒代化和香叶化。在这项研究中，我们探索了 SelU 介导的 tRNA 反密码子茎环（ASL）的体外香叶化过程，并表明当 U 突变为 A 时（ASL-tRNA 为 ASL-tRNA ），香叶化活性被废除。通过检查 SelU 辅助因子香叶基焦磷酸（gePP）及其类似物，我们发现只有香叶基，而不是具有较短或较长萜烯链的二甲基丙烯基和法呢基焦磷酸，可以掺入 ASL。SelU 末端分析中的 tRNA 香叶化程度遵循 ASL ASL >ASL 的顺序。这些发现为 SelU 的底物识别提供了一个假设的机制，并揭示了可能影响其酶活性的关键因素。鉴于 SelU 在细菌翻译系统中发挥着重要作用，抑制这种酶并靶向其香叶化和硒代化途径可能被用作开发基于 SelU 的抗生素的有前途的策略。
