Petushkov Ivan V, Aralov Andrey V, Ivanov Igor A, Baranov Mikhail S, Zatsepin Timofey S, Kulbachinskiy Andrey V
National Research Centre "Kurchatov Institute", Moscow, 123182, Russia.
Institute of Gene Biology, Russian Academy of Sciences, Moscow, 119334, Russia.
Biochemistry (Mosc). 2024 Dec;89(12):2263-2273. doi: 10.1134/S0006297924120149.
Bacterial and viral RNA polymerases are promising targets for the development of new transcription inhibitors. One of the potential blockers of RNA synthesis is 7,8-dihydro-8-oxo-1,-ethenoadenine (oxo-εA), a synthetic compound that combines two adenine modifications: 8-oxoadenine and 1,-ethenoadenine. In this study, we synthesized oxo-εA triphosphate (oxo-εATP) and showed that it could be incorporated by the RNA-dependent RNA polymerase of SARS-CoV-2 into synthesized RNA opposite template residues A and G in the presence of Mn ions. RNA polymerase incorporated oxo-εATP opposite A residues in the template DNA strand. The presence of oxo-εA instead of adenine in the template DNA strand completely stopped transcription at the modified nucleotide. At the same time, oxo-εATP did not suppress RNA synthesis by both RNA polymerases in the presence of unmodified nucleotides. Therefore, the oxo-εA modification significantly disrupts nucleotide base pairing during RNA synthesis by RNA polymerases of different classes, and the corresponding nucleotide derivatives cannot be used as potential antiviral or antibacterial transcription inhibitors.
细菌和病毒RNA聚合酶是开发新型转录抑制剂的有前景的靶点。RNA合成的潜在阻断剂之一是7,8-二氢-8-氧代-1,-乙烯腺嘌呤（氧代-εA），一种结合了两种腺嘌呤修饰的合成化合物：8-氧代腺嘌呤和1,-乙烯腺嘌呤。在本研究中，我们合成了氧代-εA三磷酸酯（氧代-εATP），并表明在锰离子存在的情况下，它可以被SARS-CoV-2的RNA依赖性RNA聚合酶掺入合成的RNA中，与模板残基A和G相对。RNA聚合酶在模板DNA链中与A残基相对处掺入氧代-εATP。模板DNA链中存在氧代-εA而非腺嘌呤会在修饰的核苷酸处完全停止转录。同时，在未修饰核苷酸存在的情况下，氧代-εATP不会抑制两种RNA聚合酶的RNA合成。因此，氧代-εA修饰显著破坏了不同类别的RNA聚合酶在RNA合成过程中的核苷酸碱基配对，相应的核苷酸衍生物不能用作潜在的抗病毒或抗菌转录抑制剂。
