Observing inhibition of the SARS-CoV-2 helicase at single-nucleotide resolution
The genome of SARS-CoV-2 encodes for a helicase (nsp13) that is essential for viral replication and highly conserved across related viruses, making it an attractive antiviral target. Here we use nanopore tweezers, a high-resolution single-molecule technique, to gain detailed insight into how nsp13 t...
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| Veröffentlicht in: | Nucleic acids research 2023-09, Vol.51 (17), p.9266-9278 |
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| creator | Marx, Sinduja K Mickolajczyk, Keith J Craig, Jonathan M Thomas, Christopher A Pfeffer, Akira M Abell, Sarah J Carrasco, Jessica D Franzi, Michaela C Huang, Jesse R Kim, Hwanhee C Brinkerhoff, Henry Kapoor, Tarun M Gundlach, Jens H Laszlo, Andrew H |
| description | The genome of SARS-CoV-2 encodes for a helicase (nsp13) that is essential for viral replication and highly conserved across related viruses, making it an attractive antiviral target. Here we use nanopore tweezers, a high-resolution single-molecule technique, to gain detailed insight into how nsp13 turns ATP-hydrolysis into directed motion along nucleic acid strands. We measured nsp13 both as it translocates along single-stranded DNA or unwinds double-stranded DNA. Our data reveal nsp13's single-nucleotide steps, translocating at ∼1000 nt/s or unwinding at ∼100 bp/s. Nanopore tweezers' high spatiotemporal resolution enables detailed kinetic analysis of nsp13 motion. As a proof-of-principle for inhibition studies, we observed nsp13's motion in the presence of the ATPase inhibitor ATPγS. We construct a detailed picture of inhibition in which ATPγS has multiple mechanisms of inhibition. The dominant mechanism of inhibition depends on the application of assisting force. This lays the groundwork for future single-molecule inhibition studies with viral helicases. |
| doi_str_mv | 10.1093/nar/gkad660 |
| format | Article |
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This lays the groundwork for future single-molecule inhibition studies with viral helicases.</description><identifier>ISSN: 0305-1048</identifier><identifier>EISSN: 1362-4962</identifier><identifier>DOI: 10.1093/nar/gkad660</identifier><identifier>PMID: 37560916</identifier><language>eng</language><publisher>England: Oxford University Press</publisher><subject>Nucleic Acid Enzymes</subject><ispartof>Nucleic acids research, 2023-09, Vol.51 (17), p.9266-9278</ispartof><rights>The Author(s) 2023. Published by Oxford University Press on behalf of Nucleic Acids Research.</rights><rights>The Author(s) 2023. 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| subjects | Nucleic Acid Enzymes |
| title | Observing inhibition of the SARS-CoV-2 helicase at single-nucleotide resolution |
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