skip to main content
OSTI.GOV title logo U.S. Department of Energy
Office of Scientific and Technical Information

Title: Activity profiling and crystal structures of inhibitor-bound SARS-CoV-2 papain-like protease: A framework for anti–COVID-19 drug design

Journal Article · · Science Advances
ORCiD logo [1];  [2]; ORCiD logo [1];  [3]; ORCiD logo [2]; ORCiD logo [4]; ORCiD logo [5];  [3]; ORCiD logo [6]; ORCiD logo [7];  [2]
  1. Wroclaw Univ. of Science and Technology, Wroclaw (Poland)
  2. Medical Univ. of South Carolina, Charleston, SC (United States); Univ. of Texas Health Science Center, San Antonio, TX (United States)
  3. New York Univ. (NYU), NY (United States)
  4. Sanford Burnham Prebys Medical Discovery Inst., La Jolla, CA (United States)
  5. UbiQ Bio B.V., Amsterdam (Netherlands)
  6. Independent Consultant; Arvinas Inc., New Haven, CT (United States)
  7. Wroclaw Univ. of Science and Technology, Wroclaw (Poland); Sanford Burnham Prebys Medical Discovery Inst., La Jolla, CA (United States)
+ Show Author Affiliations

Viral papain-like cysteine protease (PLpro, NSP3) is essential for SARS-CoV-2 replication and represents a promising target for the development of antiviral drugs. Here, we used a combinatorial substrate library and performed comprehensive activity profiling of SARS-CoV-2 PLpro. On the scaffold of the best hits from positional scanning, we designed optimal fluorogenic substrates and irreversible inhibitors with a high degree of selectivity for SARS PLpro. We determined crystal structures of two of these inhibitors in complex with SARS-CoV-2 PLpro that reveals their inhibitory mechanisms and provides a molecular basis for the observed substrate specificity profiles. Last, we demonstrate that SARS-CoV-2 PLpro harbors deISGylating activity similar to SARSCoV-1 PLpro but its ability to hydrolyze K48-linked Ub chains is diminished, which our sequence and structure analysis provides a basis for. Together, this work has revealed the molecular rules governing PLpro substrate specificity and provides a framework for development of inhibitors with potential therapeutic value or drug repurposing.

Research Organization:
Argonne National Lab. (ANL), Argonne, IL (United States). Advanced Photon Source (APS)
Sponsoring Organization:
Nationsl Science Center; National Institutes of Health (NIH)
Grant/Contract Number:
2015/17/N/ST5/03072; P30 GM124165; S10 RR027139-01; RR200030; R01 GM115568; ES025166; GM099040
OSTI ID:
1682337
Journal Information:
Science Advances, Vol. 6, Issue 42; ISSN 2375-2548
Publisher:
AAASCopyright Statement
Country of Publication:
United States
Language:
ENGLISH

References (44)

Positional-scanning fluorigenic substrate libraries reveal unexpected specificity determinants of DUBs (deubiquitinating enzymes) journal October 2008
Structural basis of the specificity of USP18 toward ISG15 journal February 2017
Isolation of a Novel Coronavirus from a Man with Pneumonia in Saudi Arabia journal November 2012
Severe Acute Respiratory Syndrome Coronavirus Papain-Like Protease Ubiquitin-Like Domain and Catalytic Domain Regulate Antagonism of IRF3 and NF-κB Signaling journal April 2009
Activity, Specificity, and Probe Design for the Smallpox Virus Protease K7L journal September 2012
Epidemiology and cause of severe acute respiratory syndrome (SARS) in Guangdong, People's Republic of China, in February, 2003 journal October 2003
A novel coronavirus outbreak of global health concern journal February 2020
Structural Basis for the Ubiquitin-Linkage Specificity and deISGylating Activity of SARS-CoV Papain-Like Protease journal May 2014
SARS and MERS: recent insights into emerging coronaviruses journal June 2016
Expedient Solid-Phase Synthesis of Fluorogenic Protease Substrates Using the 7-Amino-4-carbamoylmethylcoumarin (ACC) Fluorophore journal February 2002
Selective Substrates and Activity-Based Probes for Imaging of the Human Constitutive 20S Proteasome in Cells and Blood Samples journal December 2017
Clinical features of patients infected with 2019 novel coronavirus in Wuhan, China journal February 2020
A Native Chemical Ligation Handle that Enables the Synthesis of Advanced Activity-Based Probes: Diubiquitin as a Case Study journal March 2014
Identification of Severe Acute Respiratory Syndrome Coronavirus Replicase Products and Characterization of Papain-Like Protease Activity journal December 2004
Ubiquitin-Based Probes Prepared by Total Synthesis To Profile the Activity of Deubiquitinating Enzymes journal September 2012
The Papain-Like Protease from the Severe Acute Respiratory Syndrome Coronavirus Is a Deubiquitinating Enzyme journal November 2005
Disulfiram can inhibit MERS and SARS coronavirus papain-like proteases via different modes journal February 2018
SARS hCoV papain-like protease is a unique Lys48 linkage-specific di-distributive deubiquitinating enzyme journal May 2015
Targeting proteases: successes, failures and future prospects journal September 2006
Emerging principles in protease-based drug discovery journal September 2010
Engineered unnatural ubiquitin for optimal detection of deubiquitinating enzymes journal January 2020
MERS-CoV papain-like protease has deISGylating and deubiquitinating activities journal February 2014
Non-hydrolyzable Diubiquitin Probes Reveal Linkage-Specific Reactivity of Deubiquitylating Enzymes Mediated by S2 Pockets journal April 2016
From SARS to MERS: crystallographic studies on coronaviral proteases enable antiviral drug design journal August 2014
Regulation of IRF-3-dependent Innate Immunity by the Papain-like Protease Domain of the Severe Acute Respiratory Syndrome Coronavirus journal August 2007
Chemical Synthesis of Ubiquitin, Ubiquitin-Based Probes, and Diubiquitin journal November 2010
Selectivity in ISG15 and ubiquitin recognition by the SARS coronavirus papain-like protease journal October 2007
Profiling of flaviviral NS2B-NS3 protease specificity provides a structural basis for the development of selective chemical tools that differentiate Dengue from Zika and West Nile viruses journal March 2020
A noncovalent class of papain-like protease/deubiquitinase inhibitors blocks SARS virus replication journal October 2008
Emerging challenges in the design of selective substrates, inhibitors and activity-based probes for indistinguishable proteases journal January 2017
Coot model-building tools for molecular graphics journal November 2004
The SARS-coronavirus papain-like protease: Structure, function and inhibition by designed antiviral compounds journal March 2015
The Ubiquitin-Conjugating System: Multiple Roles in Viral Replication and Infection journal May 2014
Extended substrate specificity and first potent irreversible inhibitor/activity-based probe design for Zika virus NS2B-NS3 protease journal March 2017
PHENIX: a comprehensive Python-based system for macromolecular structure solution journal January 2010
On Terminal Alkynes That Can React with Active-Site Cysteine Nucleophiles in Proteases journal February 2013
Characterization and Noncovalent Inhibition of the Deubiquitinase and deISGylase Activity of SARS-CoV-2 Papain-Like Protease journal May 2020
Reactive-site-centric chemoproteomics identifies a distinct class of deubiquitinase enzymes journal March 2018
Recognition of Lys48-Linked Di-ubiquitin and Deubiquitinating Activities of the SARS Coronavirus Papain-like Protease journal May 2016
Deubiquitinating and Interferon Antagonism Activities of Coronavirus Papain-Like Proteases journal February 2010
The Papain-Like Protease of Severe Acute Respiratory Syndrome Coronavirus Has Deubiquitinating Activity journal November 2005
Papain-Like Protease 2 (PLP2) from Severe Acute Respiratory Syndrome Coronavirus (SARS-CoV):  Expression, Purification, Characterization, and Inhibition † journal August 2005
Synthesis of a HyCoSuL peptide substrate library to dissect protease substrate specificity journal September 2017
Dramatic Effect of Boron-Based Lewis Acids in Cross-Metathesis Reactions journal January 2005

Similar Records

Related Subjects

59 BASIC BIOLOGICAL SCIENCES