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Understanding chemical reactivity for homo- and heterobifunctional protein cross-linking agents.
Journal article

Understanding chemical reactivity for homo- and heterobifunctional protein cross-linking agents.

  • Chen F Department of Chemistry and Applied Biosciences, ETH Zürich, CH-8093, Zürich, Switzerland.
  • Nielsen S
  • Zenobi R
  • 2013-07-09
Published in:
  • Journal of mass spectrometry : JMS. - 2013
2-pyridyldithiol
MALDI mass spectrometry
N-hydroxysuccinimide (NHS) esters
amine-to-sulfhydryl cross-links
chemical cross-linking
maleimides
Cross-Linking Reagents
Glyceraldehyde-3-Phosphate Dehydrogenases
Maleimides
Models, Molecular
Protein Subunits
Proteins
Ribonucleases
Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
Succinimides
Sulfhydryl Compounds
English Chemical cross-linking, combined with mass spectrometry, has been applied to map three-dimensional protein structures and protein-protein interactions. Proper choice of the cross-linking agent, including its reactive groups and spacer arm length, is of great importance. However, studies to understand the details of reactivity of the chemical cross-linkers with proteins are quite sparse. In this study, we investigated chemical cross-linking from the aspects of the protein structures and the cross-linking reagents involved, by using two structurally well-known proteins, glyceraldehyde 3-phosohate dehydrogenase and ribonuclease S. Chemical cross-linking reactivity was compared using a series of homo- and hetero-bifunctional cross-linkers, including bis(sulfosuccinimidyl) suberate, dissuccinimidyl suberate, bis(succinimidyl) penta (ethylene glycol), bis(succinimidyl) nona (ethylene glycol), m-maleimidobenzoyl-N-hydroxysulfosuccinimide ester, 2-pyridyldithiol-tetraoxaoctatriacontane-N-hydrosuccinimide and succinimidyl-[(N-maleimidopropionamido)-tetracosaethyleneglycol]ester. The protein structure itself, especially the distances between target amino acid residues, was found to be a determining factor for the cross-linking efficiency. Moreover, the reactive groups of the chemical cross-linker also play an important role; a higher cross-linking reaction efficiency was found for maleimides compared to 2-pyrimidyldithiols. The reaction between maleimides and sulfhydryl groups is more favorable than that between N-hydroxysuccinimide esters and amine groups, although cysteine residues are less abundant in proteins compared to lysine residues.
Language
  • English
Open access status
closed
Identifiers
Persistent URL
https://folia.unifr.ch/global/documents/88826
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