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呼吸道合胞病毒聚合酶在 RNA 合成中的模板变异分析。

Analysis of Template Variations on RNA Synthesis by Respiratory Syncytial Virus Polymerase.

机构信息

Department of Biochemistry, Emory University School of Medicine, Atlanta, GA 30322, USA.

出版信息

Viruses. 2022 Dec 23;15(1):47. doi: 10.3390/v15010047.

Abstract

Respiratory syncytial virus (RSV) is a significant threat to infants and elderly individuals globally. Currently, there are no effective therapies or treatments for RSV infection because of an insufficient understanding of the RSV viral machinery. In this study, we investigated the effects of the template variations on RNA synthesis by the RSV polymerase through in vitro RNA synthesis assays. We confirmed the previously reported back-priming activity of the RSV polymerase, which is likely due to the secondary structure of the RNA template. We found that the expansion of the hairpin loop size of the RNA template abolishes the RSV polymerase back-priming activity. At the same time, it seemingly does not affect the de novo RNA synthesis activities of the RSV polymerase. Interestingly, our results show that the RSV polymerase also has a new primer-based terminal extension activity that adds nucleotides to the template and primer in a nonspecific manner. We also mapped the impact of the RNA 5' chemical group on its mobility in a urea-denaturing RNA gel shift assay. Overall, these results enhance our knowledge about the RNA synthesis processes of the RSV polymerase and may guide future therapeutic efforts to develop effective antiviral drugs for RSV treatment.

摘要

呼吸道合胞病毒(RSV)是全球婴儿和老年人的重大威胁。由于对 RSV 病毒机制的了解不足,目前尚无有效的治疗方法或药物来治疗 RSV 感染。在这项研究中,我们通过体外 RNA 合成测定研究了 RSV 聚合酶模板变化对 RNA 合成的影响。我们证实了 RSV 聚合酶先前报道的后引发活性,这可能是由于 RNA 模板的二级结构。我们发现,RNA 模板发夹环大小的扩展会破坏 RSV 聚合酶的后引发活性。同时,它似乎不会影响 RSV 聚合酶的从头 RNA 合成活性。有趣的是,我们的结果表明,RSV 聚合酶还具有新的基于引物的末端延伸活性,以非特异性的方式将核苷酸添加到模板和引物上。我们还通过尿素变性 RNA 凝胶迁移分析测定研究了 RNA 5' 化学基团对其迁移率的影响。总的来说,这些结果增强了我们对 RSV 聚合酶 RNA 合成过程的认识,并可能为未来开发有效治疗 RSV 的抗病毒药物的治疗方法提供指导。

https://cdn.ncbi.nlm.nih.gov/pmc/blobs/0afb/9863079/9383cc45b691/viruses-15-00047-g001.jpg

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