Characterization and Noncovalent Inhibition of the Deubiquitinase and deISGylase Activity of SARS-CoV-2 Papain-Like Protease

ACS Infect Dis. 2020 Aug 14;6(8):2099-2109. doi: 10.1021/acsinfecdis.0c00168. Epub 2020 Jun 4.


Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), the causative agent for COVID-19, is a novel human betacoronavirus that is rapidly spreading worldwide. The outbreak currently includes over 3.7 million cases and 260,000 fatalities. As a betacoronavirus, SARS-CoV-2 encodes for a papain-like protease (PLpro) that is likely responsible for cleavage of the coronavirus (CoV) viral polypeptide. The PLpro is also responsible for suppression of host innate immune responses by virtue of its ability to reverse host ubiquitination and ISGylation events. Here, the biochemical activity of SARS-CoV-2 PLpro against ubiquitin (Ub) and interferon-stimulated gene product 15 (ISG15) substrates is evaluated, revealing that the protease has a marked reduction in its ability to process K48 linked Ub substrates compared to its counterpart in SARS-CoV. Additionally, its substrate activity more closely mirrors that of the PLpro from the Middle East respiratory syndrome coronavirus and prefers ISG15s from certain species including humans. Additionally, naphthalene based PLpro inhibitors are shown to be effective at halting SARS-CoV-2 PLpro activity as well as SARS-CoV-2 replication.

Keywords: COVID-19; ISG5; PLpro; coronavirus; severe acute respiratory syndrome 2; ubiquitin.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Amino Acid Sequence
  • Animals
  • Betacoronavirus / enzymology*
  • Binding Sites
  • COVID-19
  • Chlorocebus aethiops
  • Coronavirus 3C Proteases
  • Coronavirus Infections / virology*
  • Cysteine Endopeptidases / chemistry
  • Cysteine Endopeptidases / metabolism*
  • Cytokines / antagonists & inhibitors
  • Cytokines / chemistry
  • Cytokines / metabolism*
  • Deubiquitinating Enzymes / antagonists & inhibitors*
  • Humans
  • Naphthalenes / pharmacology
  • Pandemics
  • Pneumonia, Viral / virology*
  • Protein Binding
  • Protein Conformation
  • SARS-CoV-2
  • Substrate Specificity
  • Ubiquitin / metabolism*
  • Ubiquitins / antagonists & inhibitors
  • Ubiquitins / chemistry
  • Ubiquitins / metabolism*
  • Vero Cells
  • Viral Nonstructural Proteins / antagonists & inhibitors*
  • Viral Nonstructural Proteins / chemistry
  • Viral Nonstructural Proteins / metabolism*
  • Virus Replication / drug effects


  • Cytokines
  • Naphthalenes
  • Ubiquitin
  • Ubiquitins
  • Viral Nonstructural Proteins
  • naphthalene
  • ISG15 protein, human
  • Deubiquitinating Enzymes
  • Cysteine Endopeptidases
  • Coronavirus 3C Proteases