Title: Structural insights into the substrate specificity of the endonuclease activity of the influenza virus cap-snatching mechanism

Abstract

Abstract The endonuclease activity within the influenza virus cap-snatching process is a proven therapeutic target. The anti-influenza drug baloxavir is highly effective, but is associated with resistance mutations that threaten its clinical efficacy. The endonuclease resides within the N-terminal domain of the PA subunit (PAN) of the influenza RNA dependent RNA polymerase, and we report here complexes of PAN with RNA and DNA oligonucleotides to understand its specificity and the structural basis of baloxavir resistance mutations. The RNA and DNA oligonucleotides bind within the substrate binding groove of PAN in a similar fashion, explaining the ability of the enzyme to cleave both substrates. The individual nucleotides occupy adjacent conserved pockets that flank the two-metal active site. However, the 2′ OH of the RNA ribose moieties engage in additional interactions that appear to optimize the binding and cleavage efficiency for the natural substrate. The major baloxavir resistance mutation at position 38 is at the core of the substrate binding site, but structural studies and modeling suggest that it maintains the necessary virus fitness via compensating interactions with RNA. These studies will facilitate the development of new influenza therapeutics that spatially match the substrate and are less likely to elicit resistance mutations.

Authors:

Kumar, Gyanendra ^[1]; Cuypers, Maxime  ^[1];

• Search DOE PAGES for author "Cuypers, Maxime"
• Search DOE PAGES for ORCID "0000-0002-8826-2317"

Webby, Richard R. ^[2]; Webb, Thomas R. ^[3]; White, Stephen W.  ^[1]

• Search DOE PAGES for author "White, Stephen W."
• Search DOE PAGES for ORCID "0000-0001-8188-5944"

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+ Show Author Affiliations

1. Department of Structural Biology, Memphis, TN 38105, USA
2. Department of Infectious Diseases, St. Jude Children's Research Hospital, Memphis, TN 38105, USA
3. Wildflower Biopharma, Inc., 8650 Genesee Ave. STE 214, San Diego, CA 92122, USA, Department of Chemistry and Biochemistry, University of California, Santa Cruz, Santa Cruz, CA 95064, USA

Publication Date:

Tue Jan 19 00:00:00 EST 2021

Research Org.:

Argonne National Lab. (ANL), Argonne, IL (United States). Advanced Photon Source (APS)

Sponsoring Org.:

USDOE Office of Science (SC); National Institutes of Health (NIH)

OSTI Identifier:

1761220

Alternate Identifier(s):

OSTI ID: 1772757

Grant/Contract Number:  

W-31-109-Eng-38; AI098757

Resource Type:

Published Article

Journal Name:

Nucleic Acids Research

Additional Journal Information:

Journal Name: Nucleic Acids Research Journal Volume: 49 Journal Issue: 3; Journal ID: ISSN 0305-1048

Publisher:

Oxford University Press

Country of Publication:

United Kingdom

Language:

English

Subject:

59 BASIC BIOLOGICAL SCIENCES