Substrate Specificity and Kinetics of RNA Hydrolysis by SARS-CoV-2 NSP10/14 Exonuclease

Abstract

Severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2), the virus that causes COVID-19, continues to evolve resistance to vaccines and existing antiviral therapies at an alarming rate, increasing the need for new direct-acting antiviral drugs. Despite significant advances in our fundamental understanding of the kinetics and mechanism of viral RNA replication, there are still open questions regarding how the proofreading exonuclease (NSP10/NSP14 complex) contributes to replication fidelity and resistance to nucleoside analogs. Through single turnover kinetic analysis, we show that the preferred substrate for the exonuclease is double-stranded RNA without any mismatches. Double-stranded RNA containing a 3′-terminal remdesivir was hydrolyzed at a rate similar to a correctly base-paired cognate nucleotide. Surprisingly, single-stranded RNA or duplex RNA containing a 3′-terminal mismatch was hydrolyzed at rates 125-and 45-fold slower, respectively, compared to the correctly base-paired double-stranded RNA. These results define the substrate specificity and rate of removal of remdesivir for the exonuclease and outline rigorous kinetic assays that could help in finding nextgeneration exonuclease inhibitors or nucleoside analogs that are able to evade excision. These results also raise important questions about the role of the polymerase/exonuclease complex in proofreading during viral replication. Addressing these questions through rigorous kinetic analysis will facilitate the search for desperately needed antiviral drugs to combat COVID-19.

Risk factors for hyperglycemia in COVID‐19 patients treated with remdesivir

“…1 Furthermore, remdesivir's efficacy against various variants, including sublineages BQ.1.1 and XBB, has sustained its usage for over two-anda-half years. [2][3][4] Additionally, a meta-analysis revealed an exceptionally low pooled COVID-19 mortality rate associated with its use, and another study demonstrated its potential to reduce mortality among COVID-19 patients requiring supplemental oxygen therapy. 5,6 Despite its clinical benefits, the administration of remdesivir has been associated with the development of hyperglycemia, a condition characterized by elevated blood glucose levels both during and after therapy.…”

“… 1 Furthermore, remdesivir's efficacy against various variants, including sublineages BQ.1.1 and XBB, has sustained its usage for over two‐and‐a‐half years. 2 , 3 , 4 Additionally, a meta‐analysis revealed an exceptionally low pooled COVID‐19 mortality rate associated with its use, and another study demonstrated its potential to reduce mortality among COVID‐19 patients requiring supplemental oxygen therapy. 5 , 6 …”