Abstract
The novel Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2 or COVID-19) has caused a global pandemic. The SARS-CoV-2 RNA genome is replicated by a conserved “core” replication-transcription complex (RTC) containing an error-prone RNA-dependent RNA polymerase holoenzyme (holo-RdRp, nsp12-nsp7-nsp8) and a RNA proofreading nuclease (nsp14-nsp10). Although structures and functions of SARS-CoV-2 holo-RdRp have been extensively studied and ribonucleotide-analog inhibitors, such as Remdesivir, have been treated for COVID-19 patients, the substrate and nucleotide specificity of SARS-CoV-2 holo-RdRp remain unknown. Here, our biochemical analysis of SARS-CoV-2 holo-RdRp reveals that it has a robust DNA-dependent RNA polymerase activity, in addition to its intrinsic RNA-dependent RNA polymerase activity. Strikingly, SARS-CoV-2 holo-RdRp fully extends RNAs with a low-fidelity even when only ATP and pyrimidine nucleotides, in particular CTP, are provided. This ATP-dependent error-prone ribonucleotide incorporation by SARS-CoV-2 holo-RdRp resists excision by the RNA proofreading nuclease in vitro. Our collective results suggest that a physiological concentration of ATP likely contributes to promoting the error-prone incorporation of ribonucleotides and ribonucleotide-analogs by SARS-CoV-2 holo-RdRp and provide a useful foundation to develop ribonucleotide analogs as an effective therapeutic strategy to combat coronavirus-mediated outbreak.
Keywords: COVID-19; RNA proofreading; RNA replication-Transcription complex; RNA-Dependent RNA polymerase; SARS-CoV-2.
【저자키워드】 COVID-19, SARS-CoV-2., RNA-dependent RNA polymerase, RNA replication-Transcription complex, RNA proofreading, 【초록키워드】 Structure, SARS-CoV-2, Genome, in vitro, inhibitors, coronavirus 2, RNA, global pandemic, specificity, outbreak, SARS-CoV-2 RNA, ATP, RNA polymerase, respiratory, therapeutic strategy, function, COVID-19 patients, nucleotide, Proofreading, Concentration, Analysis, nucleotides, physiological, replication-transcription complex, biochemical, ribonucleotide, effective, robust, intrinsic, develop, caused, conserved, addition, provided, treated, contribute, replicated, reveal, RTC, 【제목키워드】 ATP, RNA polymerase, ENhance, the SARS-CoV-2,