Force-dependent stimulation of RNA unwinding by SARS-CoV-2 nsp13 helicase |
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| Authors: | Keith J. Mickolajczyk Patrick M.M. Shelton Michael Grasso Xiaocong Cao Sara E. Warrington Amol Aher Shixin Liu Tarun M. Kapoor |
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| Affiliation: | 1. Laboratory of Chemistry and Cell Biology, The Rockefeller University, New York, New York;2. Laboratory of Nanoscale Biophysics and Biochemistry, The Rockefeller University, New York, New York;3. Laboratory of Structural Immunology, University of Science and Technology of China, Hefei, Anhui, China |
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| Abstract: | The superfamily 1 helicase nonstructural protein 13 (nsp13) is required for SARS-CoV-2 replication. The mechanism and regulation of nsp13 has not been explored at the single-molecule level. Specifically, force-dependent unwinding experiments have yet to be performed for any coronavirus helicase. Here, using optical tweezers, we find that nsp13 unwinding frequency, processivity, and velocity increase substantially when a destabilizing force is applied to the RNA substrate. These results, along with bulk assays, depict nsp13 as an intrinsically weak helicase that can be activated >50-fold by piconewton forces. Such force-dependent behavior contrasts the known behavior of other viral monomeric helicases, such as hepatitis C virus NS3, and instead draws stronger parallels to ring-shaped helicases. Our findings suggest that mechanoregulation, which may be provided by a directly bound RNA-dependent RNA polymerase, enables on-demand helicase activity on the relevant polynucleotide substrate during viral replication. |
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