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10.1128/AAC.01508-20

http://scihub22266oqcxt.onion/10.1128/AAC.01508-20
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33122171!7927875!33122171
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suck abstract from ncbi


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pmid33122171      Antimicrob+Agents+Chemother 2020 ; 65 (1): ä
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  • Development of a Simple In Vitro Assay To Identify and Evaluate Nucleotide Analogs against SARS-CoV-2 RNA-Dependent RNA Polymerase #MMPMID33122171
  • Lu G; Zhang X; Zheng W; Sun J; Hua L; Xu L; Chu XJ; Ding S; Xiong W
  • Antimicrob Agents Chemother 2020[Dec]; 65 (1): ä PMID33122171show ga
  • Nucleotide analogs targeting viral RNA polymerase have been proved to be an effective strategy for antiviral treatment and are promising antiviral drugs to combat the current severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) pandemic. In this study, we developed a robust in vitro nonradioactive primer extension assay to quantitatively evaluate the efficiency of incorporation of nucleotide analogs by SARS-CoV-2 RNA-dependent RNA polymerase (RdRp). Our results show that many nucleotide analogs can be incorporated into RNA by SARS-CoV-2 RdRp and that the incorporation of some of them leads to chain termination. The discrimination values of nucleotide analogs over those of natural nucleotides were measured to evaluate the incorporation efficiency of nucleotide analog by SARS-CoV-2 RdRp. In agreement with the data published in the literature, we found that the incorporation efficiency of remdesivir-TP is higher than that of ATP and incorporation of remdesivir-TP caused delayed chain termination, which can be overcome by higher concentrations of the next nucleotide to be incorporated. Our data also showed that the delayed chain termination pattern caused by remdesivir-TP incorporation is different for different template sequences. Multiple incorporations of remdesivir-TP caused chain termination under our assay conditions. Incorporation of sofosbuvir-TP is very low, suggesting that sofosbuvir may not be very effective in treating SARS-CoV-2 infection. As a comparison, 2'-C-methyl-GTP can be incorporated into RNA efficiently, and the derivative of 2'-C-methyl-GTP may have therapeutic application in treating SARS-CoV-2 infection. This report provides a simple screening method that should be useful for evaluating nucleotide-based drugs targeting SARS-CoV-2 RdRp and for studying the mechanism of action of selected nucleotide analogs.
  • |Adenosine Monophosphate/analogs & derivatives/chemistry/genetics/pharmacology[MESH]
  • |Alanine/analogs & derivatives/chemistry/genetics/pharmacology[MESH]
  • |Antiviral Agents/chemistry/*pharmacology[MESH]
  • |Coronavirus RNA-Dependent RNA Polymerase/antagonists & inhibitors/*genetics/metabolism[MESH]
  • |Drug Evaluation, Preclinical/*methods[MESH]
  • |Nucleotides/chemistry/*pharmacology[MESH]
  • |RNA[MESH]
  • |RNA, Viral/biosynthesis[MESH]


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