Proteome-wide covalent ligand discovery in native biological systems

Nature. 2016 Jun 23;534(7608):570-4. doi: 10.1038/nature18002. Epub 2016 Jun 15.

Abstract

Small molecules are powerful tools for investigating protein function and can serve as leads for new therapeutics. Most human proteins, however, lack small-molecule ligands, and entire protein classes are considered 'undruggable'. Fragment-based ligand discovery can identify small-molecule probes for proteins that have proven difficult to target using high-throughput screening of complex compound libraries. Although reversibly binding ligands are commonly pursued, covalent fragments provide an alternative route to small-molecule probes, including those that can access regions of proteins that are difficult to target through binding affinity alone. Here we report a quantitative analysis of cysteine-reactive small-molecule fragments screened against thousands of proteins in human proteomes and cells. Covalent ligands were identified for >700 cysteines found in both druggable proteins and proteins deficient in chemical probes, including transcription factors, adaptor/scaffolding proteins, and uncharacterized proteins. Among the atypical ligand-protein interactions discovered were compounds that react preferentially with pro- (inactive) caspases. We used these ligands to distinguish extrinsic apoptosis pathways in human cell lines versus primary human T cells, showing that the former is largely mediated by caspase-8 while the latter depends on both caspase-8 and -10. Fragment-based covalent ligand discovery provides a greatly expanded portrait of the ligandable proteome and furnishes compounds that can illuminate protein functions in native biological systems.

Publication types

  • Research Support, N.I.H., Extramural

MeSH terms

  • Apoptosis
  • Caspase 10 / chemistry
  • Caspase 10 / metabolism
  • Caspase 8 / chemistry
  • Caspase 8 / metabolism
  • Cells, Cultured
  • Cysteine / metabolism*
  • Drug Evaluation, Preclinical / methods*
  • Enzyme Precursors / chemistry
  • Enzyme Precursors / metabolism
  • Humans
  • Ligands
  • Peptide Fragments / chemistry
  • Peptide Fragments / metabolism
  • Proteome / chemistry*
  • Proteome / metabolism*
  • Small Molecule Libraries / metabolism*
  • Small Molecule Libraries / pharmacology*
  • T-Lymphocytes / chemistry
  • T-Lymphocytes / metabolism*
  • Transcription Factors / chemistry
  • Transcription Factors / metabolism

Substances

  • Enzyme Precursors
  • Ligands
  • Peptide Fragments
  • Proteome
  • Small Molecule Libraries
  • Transcription Factors
  • CASP8 protein, human
  • Caspase 10
  • Caspase 8
  • CASP10 protein, human
  • Cysteine