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Crystal structure of the Middle East respiratory syndrome coronavirus (MERS-CoV) papain-like protease bound to ubiquitin facilitates targeted disruption of deubiquitinating activity to demonstrate its role in innate immune suppression.
J Biol Chem. 2014 Dec 12; 289(50):34667-82.JB

Abstract

Middle East respiratory syndrome coronavirus (MERS-CoV) is a newly emerging human pathogen that was first isolated in 2012. MERS-CoV replication depends in part on a virus-encoded papain-like protease (PL(pro)) that cleaves the viral replicase polyproteins at three sites releasing non-structural protein 1 (nsp1), nsp2, and nsp3. In addition to this replicative function, MERS-CoV PL(pro) was recently shown to be a deubiquitinating enzyme (DUB) and to possess deISGylating activity, as previously reported for other coronaviral PL(pro) domains, including that of severe acute respiratory syndrome coronavirus. These activities have been suggested to suppress host antiviral responses during infection. To understand the molecular basis for ubiquitin (Ub) recognition and deconjugation by MERS-CoV PL(pro), we determined its crystal structure in complex with Ub. Guided by this structure, mutations were introduced into PL(pro) to specifically disrupt Ub binding without affecting viral polyprotein cleavage, as determined using an in trans nsp3↓4 cleavage assay. Having developed a strategy to selectively disable PL(pro) DUB activity, we were able to specifically examine the effects of this activity on the innate immune response. Whereas the wild-type PL(pro) domain was found to suppress IFN-β promoter activation, PL(pro) variants specifically lacking DUB activity were no longer able to do so. These findings directly implicate the DUB function of PL(pro), and not its proteolytic activity per se, in the inhibition of IFN-β promoter activity. The ability to decouple the DUB activity of PL(pro) from its role in viral polyprotein processing now provides an approach to further dissect the role(s) of PL(pro) as a viral DUB during MERS-CoV infection.

Authors+Show Affiliations

From the Department of Microbiology, University of Manitoba, Winnipeg, Manitoba R3T 2N2, Canada and.the Molecular Virology Laboratory, Department of Medical Microbiology, Leiden University Medical Center, 2333 ZA Leiden, The Netherlands.From the Department of Microbiology, University of Manitoba, Winnipeg, Manitoba R3T 2N2, Canada and.the Molecular Virology Laboratory, Department of Medical Microbiology, Leiden University Medical Center, 2333 ZA Leiden, The Netherlands.the Molecular Virology Laboratory, Department of Medical Microbiology, Leiden University Medical Center, 2333 ZA Leiden, The Netherlands.the Molecular Virology Laboratory, Department of Medical Microbiology, Leiden University Medical Center, 2333 ZA Leiden, The Netherlands.the Molecular Virology Laboratory, Department of Medical Microbiology, Leiden University Medical Center, 2333 ZA Leiden, The Netherlands.the Molecular Virology Laboratory, Department of Medical Microbiology, Leiden University Medical Center, 2333 ZA Leiden, The Netherlands.the Molecular Virology Laboratory, Department of Medical Microbiology, Leiden University Medical Center, 2333 ZA Leiden, The Netherlands m.kikkert@lumc.nl.From the Department of Microbiology, University of Manitoba, Winnipeg, Manitoba R3T 2N2, Canada and brian.mark@umanitoba.ca.

Pub Type(s)

Journal Article
Research Support, Non-U.S. Gov't

Language

eng

PubMed ID

25320088

Citation

Bailey-Elkin, Ben A., et al. "Crystal Structure of the Middle East Respiratory Syndrome Coronavirus (MERS-CoV) Papain-like Protease Bound to Ubiquitin Facilitates Targeted Disruption of Deubiquitinating Activity to Demonstrate Its Role in Innate Immune Suppression." The Journal of Biological Chemistry, vol. 289, no. 50, 2014, pp. 34667-82.
Bailey-Elkin BA, Knaap RC, Johnson GG, et al. Crystal structure of the Middle East respiratory syndrome coronavirus (MERS-CoV) papain-like protease bound to ubiquitin facilitates targeted disruption of deubiquitinating activity to demonstrate its role in innate immune suppression. J Biol Chem. 2014;289(50):34667-82.
Bailey-Elkin, B. A., Knaap, R. C., Johnson, G. G., Dalebout, T. J., Ninaber, D. K., van Kasteren, P. B., Bredenbeek, P. J., Snijder, E. J., Kikkert, M., & Mark, B. L. (2014). Crystal structure of the Middle East respiratory syndrome coronavirus (MERS-CoV) papain-like protease bound to ubiquitin facilitates targeted disruption of deubiquitinating activity to demonstrate its role in innate immune suppression. The Journal of Biological Chemistry, 289(50), 34667-82. https://doi.org/10.1074/jbc.M114.609644
Bailey-Elkin BA, et al. Crystal Structure of the Middle East Respiratory Syndrome Coronavirus (MERS-CoV) Papain-like Protease Bound to Ubiquitin Facilitates Targeted Disruption of Deubiquitinating Activity to Demonstrate Its Role in Innate Immune Suppression. J Biol Chem. 2014 Dec 12;289(50):34667-82. PubMed PMID: 25320088.
* Article titles in AMA citation format should be in sentence-case
TY - JOUR T1 - Crystal structure of the Middle East respiratory syndrome coronavirus (MERS-CoV) papain-like protease bound to ubiquitin facilitates targeted disruption of deubiquitinating activity to demonstrate its role in innate immune suppression. AU - Bailey-Elkin,Ben A, AU - Knaap,Robert C M, AU - Johnson,Garrett G, AU - Dalebout,Tim J, AU - Ninaber,Dennis K, AU - van Kasteren,Puck B, AU - Bredenbeek,Peter J, AU - Snijder,Eric J, AU - Kikkert,Marjolein, AU - Mark,Brian L, Y1 - 2014/10/15/ PY - 2014/10/17/entrez PY - 2014/10/17/pubmed PY - 2015/2/13/medline KW - Cysteine Protease KW - Deubiquitylation (Deubiquitination) KW - Innate Immunity KW - Middle East Respiratory Syndrome Coronavirus KW - PLpro KW - Structural Biology KW - Viral Immunology KW - X-ray Crystallography SP - 34667 EP - 82 JF - The Journal of biological chemistry JO - J. Biol. Chem. VL - 289 IS - 50 N2 - Middle East respiratory syndrome coronavirus (MERS-CoV) is a newly emerging human pathogen that was first isolated in 2012. MERS-CoV replication depends in part on a virus-encoded papain-like protease (PL(pro)) that cleaves the viral replicase polyproteins at three sites releasing non-structural protein 1 (nsp1), nsp2, and nsp3. In addition to this replicative function, MERS-CoV PL(pro) was recently shown to be a deubiquitinating enzyme (DUB) and to possess deISGylating activity, as previously reported for other coronaviral PL(pro) domains, including that of severe acute respiratory syndrome coronavirus. These activities have been suggested to suppress host antiviral responses during infection. To understand the molecular basis for ubiquitin (Ub) recognition and deconjugation by MERS-CoV PL(pro), we determined its crystal structure in complex with Ub. Guided by this structure, mutations were introduced into PL(pro) to specifically disrupt Ub binding without affecting viral polyprotein cleavage, as determined using an in trans nsp3↓4 cleavage assay. Having developed a strategy to selectively disable PL(pro) DUB activity, we were able to specifically examine the effects of this activity on the innate immune response. Whereas the wild-type PL(pro) domain was found to suppress IFN-β promoter activation, PL(pro) variants specifically lacking DUB activity were no longer able to do so. These findings directly implicate the DUB function of PL(pro), and not its proteolytic activity per se, in the inhibition of IFN-β promoter activity. The ability to decouple the DUB activity of PL(pro) from its role in viral polyprotein processing now provides an approach to further dissect the role(s) of PL(pro) as a viral DUB during MERS-CoV infection. SN - 1083-351X UR - https://www.unboundmedicine.com/medline/citation/25320088/Crystal_structure_of_the_Middle_East_respiratory_syndrome_coronavirus__MERS_CoV__papain_like_protease_bound_to_ubiquitin_facilitates_targeted_disruption_of_deubiquitinating_activity_to_demonstrate_its_role_in_innate_immune_suppression_ L2 - http://www.jbc.org/cgi/pmidlookup?view=long&pmid=25320088 DB - PRIME DP - Unbound Medicine ER -