Proximity Labeling for the Identification of Coronavirus-Host Protein Interactions

Methods Mol Biol. 2020:2203:187-204. doi: 10.1007/978-1-0716-0900-2_14.


Biotin-based proximity labeling circumvents major pitfalls of classical biochemical approaches to identify protein-protein interactions. It consists of enzyme-catalyzed biotin tags ubiquitously apposed on proteins located in close proximity of the labeling enzyme, followed by affinity purification and identification of biotinylated proteins by mass spectrometry. Here we outline the methods by which the molecular microenvironment of the coronavirus replicase/transcriptase complex (RTC), i.e., proteins located within a close perimeter of the RTC, can be determined by different proximity labeling approaches using BirAR118G (BioID), TurboID, and APEX2. These factors represent a molecular signature of coronavirus RTCs and likely contribute to the viral life cycle, thereby constituting attractive targets for the development of antiviral intervention strategies.

Keywords: Affinity purification; Ascorbate peroxidase APEX2; BioID; Biotin ligase; Coronavirus; Mass spectrometry; Proteomics; Proximity labeling; Replicase microenvironment; Replication transcription complex RTC; TurboID.

MeSH terms

  • Animals
  • Ascorbate Peroxidases / genetics
  • Biotinylation
  • Carbon-Nitrogen Ligases / genetics
  • Cell Line
  • Coronavirus / genetics
  • Coronavirus / pathogenicity*
  • Enzymes / genetics*
  • Enzymes / metabolism
  • Escherichia coli Proteins / genetics
  • Fluorescent Antibody Technique
  • Host-Pathogen Interactions / physiology*
  • Microorganisms, Genetically-Modified
  • Proteomics / methods*
  • Repressor Proteins / genetics
  • Viral Proteins / chemistry
  • Viral Proteins / genetics
  • Viral Proteins / metabolism*


  • Enzymes
  • Escherichia coli Proteins
  • Repressor Proteins
  • Viral Proteins
  • Ascorbate Peroxidases
  • Carbon-Nitrogen Ligases
  • birA protein, E coli