Crystal structure of the SARS-CoV-2 non-structural protein 9, Nsp9

  1. D. R. Littler
  2. B. S. Gully
  3. R. N. Colson
  4. J Rossjohn

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Abstract

Many of the proteins produced by SARS-CoV-2 have related counterparts across the Severe Acute Respiratory Syndrome (SARS-CoV) family. One such protein is non-structural protein 9 (Nsp9), which is thought to mediate both viral replication and virulence. Current understanding suggests that Nsp9 is involved in viral genomic RNA reproduction. Nsp9 is thought to bind RNA via a fold that is unique to this class of betacoronoaviruses although the molecular basis for this remains ill-defined. We sought to better characterise the SARS-CoV-2 Nsp9 protein and subsequently solved its X-ray crystal structure, in an apo-form and, unexpectedly, in a peptide-bound form with a sequence originating from a rhinoviral 3C protease sequence (LEVL). The structure of the SARS-CoV-2 Nsp9 revealed the high level of structural conservation within the Nsp9 family. The exogenous peptide binding site is close to the dimer interface and impacted on the relative juxtaposition of the monomers within the homodimer. Together we have established a protocol for the production of SARS-CoV-2 Nsp9, determined its structure and identified a peptide-binding site that may warrant further study from the perspective of understanding Nsp9 function.

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  1. ScreenIT

    SciScore for 10.1101/2020.03.28.013920: (What is this?)

    Please note, not all rigor criteria are appropriate for all manuscripts.

    Table 1: Rigor

    Institutional Review Board Statementnot detected.
    Randomizationnot detected.
    Blindingnot detected.
    Power Analysisnot detected.
    Sex as a biological variablenot detected.
    Cell Line AuthenticationAuthentication: Crystals of the His-tag samples grew with rectangular morphology in space group P4122, however if the His-tag was removed the crystals grew in space group P6122 form with hexagonal morphology.

    Table 2: Resources

    Software and Algorithms
    SentencesResources
    Subsequent rounds of manual building and refinement were performed in Coot (Casanal et al., 2019) and PHENIX (Liebschner et al., 2019).
    Coot
    suggested: (Coot, RRID:SCR_014222)
    PHENIX
    suggested: (Phenix, RRID:SCR_014224)
    Amax and KD were used as fitting parameters and nonlinear regression was performed using GraphPad Prism.
    GraphPad Prism
    suggested: (GraphPad Prism, RRID:SCR_002798)

    Results from OddPub: Thank you for sharing your data.

    Results from LimitationRecognizer: An explicit section about the limitations of the techniques employed in this study was not found. We encourage authors to address study limitations.

    Results from TrialIdentifier: No clinical trial numbers were referenced.

    Results from Barzooka: We did not find any issues relating to the usage of bar graphs.

    Results from JetFighter: We did not find any issues relating to colormaps.

    Results from rtransparent:
    • Thank you for including a conflict of interest statement. Authors are encouraged to include this statement when submitting to a journal.
    • Thank you for including a funding statement. Authors are encouraged to include this statement when submitting to a journal.
    • No protocol registration statement was detected.

    About SciScore

    SciScore is an automated tool that is designed to assist expert reviewers by finding and presenting formulaic information scattered throughout a paper in a standard, easy to digest format. SciScore checks for the presence and correctness of RRIDs (research resource identifiers), and for rigor criteria such as sex and investigator blinding. For details on the theoretical underpinning of rigor criteria and the tools shown here, including references cited, please follow this link.

  2. Version published to 10.1101/2020.03.28.013920 on bioRxiv