Nidovirus papain-like proteases: structural insight into substrate recognition and innate immune suppression
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Date
2014-03-27, 2013-02-26, 2017-05-18, 2017-06-16, 2014-12-12
Authors
Bailey-Elkin, Ben A
Journal Title
Journal ISSN
Volume Title
Publisher
Public Library of Science
United States National Academy of Sciences
Public Library of Science
Elsevier
American Society for Biochemistry and Molecular Biology
United States National Academy of Sciences
Public Library of Science
Elsevier
American Society for Biochemistry and Molecular Biology
Abstract
Nidoviruses are an order of positive-sense RNA viruses, which include the families Arteriviridae and Coronaviridae. Nidoviruses express their complement of non-structural proteins (nsps) as a single polyprotein, which is cleaved into functional domains by proteases encoded within. The equine arterivirus (EAV) and Middle East respiratory syndrome coronavirus (MERS-CoV) encode for papain-like protease domains within nsp2 and nsp3, respectively, which serve a replicative role through their polyprotein processing activities.
During infection, the host innate immune response is triggered by incoming pathogens, and engages multiple signalling pathways which are in-part dependent on the post-translational modification by ubiquitin (Ub). These signalling processes are tightly regulated by deubiquitinases (DUBs), which remove Ub from their cellular targets in order to reverse their effects.
The EAV papain-like protease 2 (PLP2) and the MERS-CoV papain-like protease (PLpro), in addition to their replicative functions, were proposed to interfere with the induction of the cellular innate immune response to infection by acting as deubiquitinating enzymes. The fact that these enzymes rely on a single active site in order to carry out both DUB and polyprotein processing activities complicates our ability to assess the role of these functions independently. Here, the crystal structures of EAV PLP2, and the MERS-CoV PLpro in covalent complex with their Ub substrates were determined, and structure-guided mutagenesis was used to selectively remove DUB activity permitting the independent study of their DUB activities. Specific mutations targeting the Ub-binding interface of these enzymes inhibited DUB activity while permitting replicative polyprotein processing. Studies using these DUB-deficient enzymes demonstrated directly their role in the down regulation of cellular innate immune responses. In an effort to further define the molecular basis for substrate recognition by these viral DUBs, their structures were determined in complex with the antiviral Ub-like molecule ISG15, permitting further structural characterization of substrate recognition.
In addition to the structural characterization of PLpro in complex with cellular substrates, the structures of PLpro in complex with novel, selective Ub-based inhibitors of the enzyme were determined. These inhibitors bound with high affinity mediated by a combination of hydrophobic and hydrogen-bonding interactions, inhibiting polyprotein cleavage, innate immune suppression and viral replication in cell culture.
Description
Keywords
innate immune evasion, ubiquitin, papain-like protease, viral deubiquitinating enzyme, Middle East Respiratory Syndrome Coronavirus, MERS, Equine arteritis virus, EAV, OTU, PLpro, X-ray Crystallography
Citation
Bailey-Elkin BA, van Kasteren PB, Snijder EJ, Kikkert M, Mark BL (2014) Viral OTU Deubiquitinases: A structural and functional comparison. PLoS Pathogens. Mar 27;10(3):e1003894
van Kasteren PB, Bailey-Elkin BA, James TW, Ninaber DK, Beugeling C, Khajehpour M, Snijder EJ, Mark BL, Kikkert M (2013) The deubiquitinase function of arterivirus papain-like protease 2 suppresses the innate immune response in infected host cells. PNAS. Feb 26;110(9):E838-47
Zhang W, Bailey-Elkin BA, Knaap RCM, Khare B, Dalebout TJ, Johnson GG, van Kasteren PB, McLeish NJ, Gu J, He W, Kikkert M, Mark BL, Sidhu SS (2017) Potent and selective inhibition of pathogenic viruses by engineered ubiquitin variants. PLoS Pathogens. May 18;13(5):e1006372
Bailey-Elkin BA, Knaap RCM, Kikkert M, Mark BL (2017) Structure and function of viral deubiquitinating enzymes. Journal of Molecular Biology. Jun 16. pii: S0022-2836(17)30310-8
Bailey-Elkin BA, Knaap RCM, Johnson GG, Dalebout TJ, Ninaber DK, van Kasteren PB, Bredenbeek PJ, Snijder EJ, Kikkert M, Mark BL (2014) Crystal structure of the MERS coronavirus papain-like protease bound to ubiquitin facilitates targeted disruption of deubiquitinating activity to demonstrate its role in innate immune suppression. Journal of Biological Chemistry. Dec 12;289(50):34667-82
van Kasteren PB, Bailey-Elkin BA, James TW, Ninaber DK, Beugeling C, Khajehpour M, Snijder EJ, Mark BL, Kikkert M (2013) The deubiquitinase function of arterivirus papain-like protease 2 suppresses the innate immune response in infected host cells. PNAS. Feb 26;110(9):E838-47
Zhang W, Bailey-Elkin BA, Knaap RCM, Khare B, Dalebout TJ, Johnson GG, van Kasteren PB, McLeish NJ, Gu J, He W, Kikkert M, Mark BL, Sidhu SS (2017) Potent and selective inhibition of pathogenic viruses by engineered ubiquitin variants. PLoS Pathogens. May 18;13(5):e1006372
Bailey-Elkin BA, Knaap RCM, Kikkert M, Mark BL (2017) Structure and function of viral deubiquitinating enzymes. Journal of Molecular Biology. Jun 16. pii: S0022-2836(17)30310-8
Bailey-Elkin BA, Knaap RCM, Johnson GG, Dalebout TJ, Ninaber DK, van Kasteren PB, Bredenbeek PJ, Snijder EJ, Kikkert M, Mark BL (2014) Crystal structure of the MERS coronavirus papain-like protease bound to ubiquitin facilitates targeted disruption of deubiquitinating activity to demonstrate its role in innate immune suppression. Journal of Biological Chemistry. Dec 12;289(50):34667-82