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Signatures and mechanisms of efficacious therapeutic ribonucleotides against SARS-CoV-2 revealed by analysis of its replicase using magnetic tweezers
Mona Seifert; Subhas C. Bera; Pauline van Nies; Robert N. Kirchdoerfer; Ashleigh Shannon; Thi-Tuyet-Nhung Le; Xiangzhi Meng; Hongjie Xia; James M. Wood; Lawrence D. Harris; Flavia S. Papini; Jamie J. Arnold; Steven C. Almo; Tyler L. Grove; Pei-Yong Shi; Yan Xiang; Bruno Canard; Martin Depken; Craig E. Cameron; David Dulin.
Affiliation
  • Mona Seifert; Junior Research Group 2, Interdisciplinary Center for Clinical Research, Friedrich-Alexander-University Erlangen-Nurnberg (FAU), Cauerstr. 3, 91058 Erlangen, Ge
  • Subhas C. Bera; Junior Research Group 2, Interdisciplinary Center for Clinical Research, Friedrich-Alexander-University Erlangen-Nurnberg (FAU), Cauerstr. 3, 91058 Erlangen, Ge
  • Pauline van Nies; Junior Research Group 2, Interdisciplinary Center for Clinical Research, Friedrich-Alexander-University Erlangen-Nurnberg (FAU), Cauerstr. 3, 91058 Erlangen, Ge
  • Robert N. Kirchdoerfer; University of Wisconsin-Madison
  • Ashleigh Shannon; Architecture et Fonction des Macromolecules Biologiques, CNRS and Aix-Marseille Universite, UMR 7257, Polytech Case 925, 13009 Marseille, France.
  • Thi-Tuyet-Nhung Le; Architecture et Fonction des Macromolecules Biologiques, CNRS and Aix-Marseille Universite, UMR 7257, Polytech Case 925, 13009 Marseille, France.
  • Xiangzhi Meng; Department of Microbiology, Immunology and Molecular Genetics, University of Texas Health Science Center at San Antonio, 7703 Floyd Curl Drive, San Antonio, TX
  • Hongjie Xia; Department of Biochemistry and Molecular Biology, University of Texas Medical Branch, Galveston, TX 77555, USA
  • James M. Wood; The Ferrier Research Institute, Victoria University of Wellington, Wellington, New Zealand
  • Lawrence D. Harris; The Ferrier Research Institute, Victoria University of Wellington, Wellington, New Zealand
  • Flavia S. Papini; Junior Research Group 2, Interdisciplinary Center for Clinical Research, Friedrich-Alexander-University Erlangen-Nurnberg (FAU), Cauerstr. 3, 91058 Erlangen, Ge
  • Jamie J. Arnold; Department of Microbiology and Immunology, University of North Carolina School of Medicine, Chapel Hill, NC 27514 USA
  • Steven C. Almo; Department of Biochemistry, Albert Einstein College of Medicine, Bronx, NY, USA Institute for Protein Innovation, Boston, MA, USA
  • Tyler L. Grove; Department of Biochemistry, Albert Einstein College of Medicine, Bronx, NY, USA Institute for Protein Innovation, Boston, MA, USA
  • Pei-Yong Shi; Department of Biochemistry and Molecular Biology, University of Texas Medical Branch, Galveston, TX 77555, USA; Institute for Human Infections and Immunity, Uni
  • Yan Xiang; Department of Microbiology, Immunology and Molecular Genetics, University of Texas Health Science Center at San Antonio, 7703 Floyd Curl Drive, San Antonio, TX
  • Bruno Canard; Architecture et Fonction des Macromolecules Biologiques, CNRS and Aix-Marseille Universite, UMR 7257, Polytech Case 925, 13009 Marseille, France.
  • Martin Depken; Department of Bionanoscience, Kavli Institute of Nanoscience, Delft University of Technology, Van der Maasweg 9, 2629 HZ Delft, The Netherlands
  • Craig E. Cameron; Department of Microbiology and Immunology, University of North Carolina School of Medicine, Chapel Hill, NC 27514 USA
  • David Dulin; Junior Research Group 2, Interdisciplinary Center for Clinical Research, Friedrich-Alexander-University Erlangen-Nurnberg (FAU), Cauerstr. 3, 91058 Erlangen, Ge
Preprint in English | bioRxiv | ID: ppbiorxiv-240325
ABSTRACT
The nucleotide analog Remdesivir (RDV) is the only FDA-approved antiviral therapy to treat infection by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). The physical basis for efficient utilization of RDV by SARS-CoV-2 polymerase is unknown. Here, we characterize the impact of RDV and other nucleotide analogs on RNA synthesis by the polymerase using a high-throughput, single-molecule, magnetic-tweezers platform. The location of the modification in the ribose or in the base dictates the catalytic pathway(s) used for its incorporation. We reveal that RDV incorporation does not terminate viral RNA synthesis, but leads the polymerase into deep backtrack, which may appear as termination in traditional ensemble assays. SARS-CoV-2 is able to evade the endogenously synthesized product of the viperin antiviral protein, ddhCTP, though the polymerase incorporates this nucleotide analog well. This experimental paradigm is essential to the discovery and development of therapeutics targeting viral polymerases. TeaserWe revise Remdesivirs mechanism of action and reveal SARS-CoV-2 ability to evade interferon-induced antiviral ddhCTP
License
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Full text: Available Collection: Preprints Database: bioRxiv Language: English Year: 2020 Document type: Preprint
Full text: Available Collection: Preprints Database: bioRxiv Language: English Year: 2020 Document type: Preprint
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