In vivo application of photocleavable protein interaction reporter technology

J Proteome Res. 2012 Feb 3;11(2):1027-41. doi: 10.1021/pr200775j. Epub 2012 Jan 9.


In vivo protein structures and protein-protein interactions are critical to the function of proteins in biological systems. As a complementary approach to traditional protein interaction identification methods, cross-linking strategies are beginning to provide additional data on protein and protein complex topological features. Previously, photocleavable protein interaction reporter (pcPIR) technology was demonstrated by cross-linking pure proteins and protein complexes and the use of ultraviolet light to cleave or release cross-linked peptides to enable identification. In the present report, the pcPIR strategy is applied to Escherichia coli cells, and in vivo protein interactions and topologies are measured. More than 1600 labeled peptides from E. coli were identified, indicating that many protein sites react with pcPIR in vivo. From those labeled sites, 53 in vivo intercross-linked peptide pairs were identified and manually validated. Approximately half of the interactions have been reported using other techniques, although detailed structures exist for very few. Three proteins or protein complexes with detailed crystallography structures are compared to the cross-linking results obtained from in vivo application of pcPIR technology.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Amino Acid Sequence
  • Binding Sites
  • Cross-Linking Reagents / chemistry
  • Cross-Linking Reagents / metabolism
  • Cross-Linking Reagents / radiation effects*
  • Escherichia coli
  • Escherichia coli Proteins / analysis
  • Escherichia coli Proteins / chemistry
  • Escherichia coli Proteins / metabolism
  • Mass Spectrometry
  • Models, Molecular
  • Molecular Sequence Data
  • Multiprotein Complexes / analysis
  • Multiprotein Complexes / chemistry*
  • Multiprotein Complexes / metabolism
  • Peptide Fragments / analysis
  • Peptide Fragments / chemistry
  • Peptide Fragments / metabolism
  • Photochemistry
  • Photolysis*
  • Protein Interaction Mapping / methods*
  • Reproducibility of Results
  • Sequence Alignment
  • Ultraviolet Rays


  • Cross-Linking Reagents
  • Escherichia coli Proteins
  • Multiprotein Complexes
  • Peptide Fragments