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Allosteric regulation and crystallographic fragment screening of SARS-CoV-2 NSP15 endoribonuclease

Godoy, Andre Schutzer ; Nakamura, Aline Minalli ; Douangamath, Alice ; Song, Yun ; Noske, Gabriela Dias ; Gawriljuk, Victor Oliveira ; Fernandes, Rafaela Sachetto ; Pereira, Humberto D.Muniz ; Oliveira, Ketllyn Irene Zagato and Fearon, Daren , et al. (2023) In Nucleic Acids Research 51(10). p.5255-5270
Abstract

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is the causative agent of coronavirus disease 2019 (COVID-19). The NSP15 endoribonuclease enzyme, known as NendoU, is highly conserved and plays a critical role in the ability of the virus to evade the immune system. NendoU is a promising target for the development of new antiviral drugs. However, the complexity of the enzyme's structure and kinetics, along with the broad range of recognition sequences and lack of structural complexes, hampers the development of inhibitors. Here, we performed enzymatic characterization of NendoU in its monomeric and hexameric form, showing that hexamers are allosteric enzymes with a positive cooperative index, and with no influence of... (More)

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is the causative agent of coronavirus disease 2019 (COVID-19). The NSP15 endoribonuclease enzyme, known as NendoU, is highly conserved and plays a critical role in the ability of the virus to evade the immune system. NendoU is a promising target for the development of new antiviral drugs. However, the complexity of the enzyme's structure and kinetics, along with the broad range of recognition sequences and lack of structural complexes, hampers the development of inhibitors. Here, we performed enzymatic characterization of NendoU in its monomeric and hexameric form, showing that hexamers are allosteric enzymes with a positive cooperative index, and with no influence of manganese on enzymatic activity. Through combining cryo-electron microscopy at different pHs, X-ray crystallography and biochemical and structural analysis, we showed that NendoU can shift between open and closed forms, which probably correspond to active and inactive states, respectively. We also explored the possibility of NendoU assembling into larger supramolecular structures and proposed a mechanism for allosteric regulation. In addition, we conducted a large fragment screening campaign against NendoU and identified several new allosteric sites that could be targeted for the development of new inhibitors. Overall, our findings provide insights into the complex structure and function of NendoU and offer new opportunities for the development of inhibitors.

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organization
publishing date
type
Contribution to journal
publication status
published
subject
in
Nucleic Acids Research
volume
51
issue
10
pages
16 pages
publisher
Oxford University Press
external identifiers
  • pmid:37115000
  • scopus:85162256992
ISSN
0305-1048
DOI
10.1093/nar/gkad314
language
English
LU publication?
yes
id
1fce2f7f-e6b9-41ef-a559-02f629d08b01
date added to LUP
2023-09-18 13:10:25
date last changed
2024-04-19 01:45:43
@article{1fce2f7f-e6b9-41ef-a559-02f629d08b01,
  abstract     = {{<p>Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is the causative agent of coronavirus disease 2019 (COVID-19). The NSP15 endoribonuclease enzyme, known as NendoU, is highly conserved and plays a critical role in the ability of the virus to evade the immune system. NendoU is a promising target for the development of new antiviral drugs. However, the complexity of the enzyme's structure and kinetics, along with the broad range of recognition sequences and lack of structural complexes, hampers the development of inhibitors. Here, we performed enzymatic characterization of NendoU in its monomeric and hexameric form, showing that hexamers are allosteric enzymes with a positive cooperative index, and with no influence of manganese on enzymatic activity. Through combining cryo-electron microscopy at different pHs, X-ray crystallography and biochemical and structural analysis, we showed that NendoU can shift between open and closed forms, which probably correspond to active and inactive states, respectively. We also explored the possibility of NendoU assembling into larger supramolecular structures and proposed a mechanism for allosteric regulation. In addition, we conducted a large fragment screening campaign against NendoU and identified several new allosteric sites that could be targeted for the development of new inhibitors. Overall, our findings provide insights into the complex structure and function of NendoU and offer new opportunities for the development of inhibitors.</p>}},
  author       = {{Godoy, Andre Schutzer and Nakamura, Aline Minalli and Douangamath, Alice and Song, Yun and Noske, Gabriela Dias and Gawriljuk, Victor Oliveira and Fernandes, Rafaela Sachetto and Pereira, Humberto D.Muniz and Oliveira, Ketllyn Irene Zagato and Fearon, Daren and Dias, Alexandre and Krojer, Tobias and Fairhead, Michael and Powell, Alisa and Dunnet, Louise and Brandao-Neto, Jose and Skyner, Rachael and Chalk, Rod and Bajusz, Dávid and Bege, Miklós and Borbás, Anikó and Keseru, György Miklós and Von Delft, Frank and Oliva, Glaucius}},
  issn         = {{0305-1048}},
  language     = {{eng}},
  number       = {{10}},
  pages        = {{5255--5270}},
  publisher    = {{Oxford University Press}},
  series       = {{Nucleic Acids Research}},
  title        = {{Allosteric regulation and crystallographic fragment screening of SARS-CoV-2 NSP15 endoribonuclease}},
  url          = {{http://dx.doi.org/10.1093/nar/gkad314}},
  doi          = {{10.1093/nar/gkad314}},
  volume       = {{51}},
  year         = {{2023}},
}