Activity-based Protein Profiling of Serine Hydrolase Superfamily Enzymes

Achintya Kumar Dolui, Mahadev Latha, Panneerselvam Vijayaraj

Research output: Contribution to journalArticlepeer-review


Activity-based protein profiling (ABPP) is a chemoproteomics platform to assess the functional state of enzymes in complex biological systems. Over the two decades, ABPP has emerged from a gel-based to gel-free platform, for in-depth proteome analysis with enhanced resolution, sensitivity for target detection, and discovery of small molecule inhibitors. The gel-free format of ABPP coupled with advanced mass spectrometry is highly sensitive and provides more comprehensive knowledge for the targeted enzyme family than the gel-based method. ABPP strategy is applied across microbe, plant, and animal models. It can be performed both in vitro and in vivo studies, and there is no limitation on sample origin. Here, we report an ultrasensitive, gel-free format of ABPP called active site peptide profiling. This protocol describes the identification of authentic functional proteins, by tagging their active sites in a native biological system. It is high throughput in nature and helps enrich even low abundance functional proteins. Since protein identification is virtually based on a single peptide, the identified peptide should be a unique peptide to identify its parent protein. It can be performed in a facile manner and offers to consolidate identification of protein targets as well as the site of probe modification. We have validated this approach using a fluorophosphonate (FP) serine hydrolase probe in the native proteome of the cereal crop Oryza sativa.

Original languageEnglish
Article numbere4356
Issue number6
StatePublished - 20 Mar 2022
Externally publishedYes


  • ABPP
  • Active site labelling
  • Peptide profiling
  • Protein identification
  • Serine hydrolase

ASJC Scopus subject areas

  • General Biochemistry, Genetics and Molecular Biology
  • General Immunology and Microbiology
  • General Neuroscience
  • Plant Science


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