Impact of Chemical Cross-Linking on Protein Structure and Function

Anal Chem. 2018 Jan 16;90(2):1104-1113. doi: 10.1021/acs.analchem.7b02863. Epub 2018 Jan 2.


Chemical cross-linking coupled with mass spectrometry is a popular technique for deriving structural information on proteins and protein complexes. Also, cross-linking has become a powerful tool for stabilizing macromolecular complexes for single-particle cryo-electron microscopy. However, an effect of cross-linking on protein structure and function should not be forgotten, and surprisingly, it has not been investigated in detail so far. Here, we used kinetic studies, mass spectrometry, and NMR spectroscopy to systematically investigate an impact of cross-linking on structure and function of human carbonic anhydrase and alcohol dehydrogenase 1 from Saccharomyces cerevisiae. We found that cross-linking induces rather local structural disturbances and the overall fold is preserved even at a higher cross-linker concentration. The results establish general experimental conditions for chemical cross-linking with minimal effect on protein structure and function.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Alcohol Dehydrogenase / chemistry*
  • Carbonic Anhydrases / chemistry*
  • Cross-Linking Reagents / chemistry*
  • Humans
  • Mass Spectrometry
  • Models, Molecular
  • Nuclear Magnetic Resonance, Biomolecular
  • Protein Conformation
  • Protein Multimerization


  • Cross-Linking Reagents
  • Alcohol Dehydrogenase
  • Carbonic Anhydrases