Room-temperature structural studies of SARS-CoV-2 protein NendoU with an X-ray free-electron laser

Rebecca J. Jernigan, Dhenugen Logeswaran, Diandra Doppler, Nirupa Nagaratnam, Mukul Sonker, Jay How Yang, Gihan Ketawala, Jose M. Martin-Garcia, Megan L. Shelby, Thomas D. Grant, Valerio Mariani, Alexandra Tolstikova, Michelle Z. Sheikh, Mimi Cho Yung, Matthew A. Coleman, Sahba Zaare, Emily K. Kaschner, Mohammad Towshif Rabbani, Reza Nazari, Michele A. ZacksBrandon Hayes, Raymond G. Sierra, Mark S. Hunter, Stella Lisova, Alexander Batyuk, Christopher Kupitz, Sebastien Boutet, Debra T. Hansen, Richard A. Kirian, Marius Schmidt, Raimund Fromme, Matthias Frank, Alexandra Ros, Julian J.L. Chen, Sabine Botha, Petra Fromme

Research output: Contribution to journalArticlepeer-review

7 Scopus citations

Abstract

NendoU from SARS-CoV-2 is responsible for the virus's ability to evade the innate immune system by cleaving the polyuridine leader sequence of antisense viral RNA. Here we report the room-temperature structure of NendoU, solved by serial femtosecond crystallography at an X-ray free-electron laser to 2.6 Å resolution. The room-temperature structure provides insight into the flexibility, dynamics, and other intrinsic properties of NendoU, with indications that the enzyme functions as an allosteric switch. Functional studies examining cleavage specificity in solution and in crystals support the uridine-purine cleavage preference, and we demonstrate that enzyme activity is fully maintained in crystal form. Optimizing the purification of NendoU and identifying suitable crystallization conditions set the benchmark for future time-resolved serial femtosecond crystallography studies. This could advance the design of antivirals with higher efficacy in treating coronaviral infections, since drugs that block allosteric conformational changes are less prone to drug resistance.

Original languageEnglish (US)
Pages (from-to)138-151.e5
JournalStructure
Volume31
Issue number2
DOIs
StatePublished - Feb 2 2023

Keywords

  • COVID-19
  • NSP15
  • NendoU
  • SARS-CoV-2
  • X-ray free-electron laser
  • serial femtosecond crystallography

ASJC Scopus subject areas

  • Structural Biology
  • Molecular Biology

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  • Room temperature structure of NSP15 Endoribonuclease from SARS CoV-2 solved using SFX.

    Jernigan, R. J. (Contributor), Logeswaran, D. (Contributor), Doppler, D. (Contributor), Nagaratnam, N. (Contributor), Sonker, M. (Contributor), Yang, J. (Contributor), Ketawala, G. (Contributor), Martin-Garcia, J. M. (Contributor), Shelby, M. L. (Contributor), Grant, T. D. (Contributor), Mariani, V. (Contributor), Tolstikova, A. (Contributor), Sheikh, M. Z. (Contributor), Yung, M. C. (Contributor), Coleman, M. A. (Contributor), Zaare, S. (Contributor), Kaschner, E. K. (Contributor), Rabbani, M. T. (Contributor), Nazari, R. (Contributor), Zacks, M. A. (Contributor), Hayes, B. (Contributor), Sierra, R. G. (Contributor), Hunter, M. S. (Contributor), Lisova, S. (Contributor), Batyuk, A. (Contributor), Kupitz, C. (Contributor), Boutet, S. (Contributor), Hansen, D. (Contributor), Kirian, R. (Contributor), Schmidt, M. (Contributor), Fromme, R. (Contributor), Frank, M. (Contributor), Ros, A. (Contributor), Chen, J. (Contributor), Botha, S. (Contributor) & Fromme, P. (Contributor), Protein Data Bank (PDB), Oct 21 2020

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