Development of a fluorescence-based method for the rapid determination of Zika virus polymerase activity and the screening of antiviral drugs

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Zika virus (ZIKV) is an emerging pathogen that has been associated with large numbers of cases of severe neurologic disease, including Guillain-Barre syndrome and microcephaly. Despite its recent establishment as a serious global public health concern there are no licensed therapeutics to control this virus. Accordingly, there is an urgent need to develop methods for the high-throughput screening of antiviral agents. We describe here a fluorescence-based method to monitor the real-time polymerization activity of Zika virus RNA-dependent RNA polymerase (RdRp). By using homopolymeric RNA template molecules, de novo RNA synthesis can be detected with a fluorescent dye, which permits the specific quantification and kinetics of double-strand RNA formation. ZIKV RdRp activity detected using this fluorescence-based assay positively correlated with traditional assays measuring the incorporation of radiolabeled nucleotides. We also validated this method as a suitable assay for the identification of ZIKV inhibitors targeting the viral polymerase using known broad-spectrum inhibitors. The assay was also successfully adapted to detect RNA polymerization activity by different RdRps, illustrated here using purified RdRps from hepatitis C virus and foot-and-mouth disease virus. The potential of fluorescence-based approaches for the enzymatic characterization of viral polymerases, as well as for high-throughput screening of antiviral drugs, are discussed.
Original languageEnglish
Article number5397
JournalScientific Reports
Volume9
Issue number1
Number of pages11
ISSN2045-2322
DOIs
Publication statusPublished - 2019
CitationsWeb of Science® Times Cited: No match on DOI
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