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

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Author(s) Chen, Fan, Nielsen, Simone, Zenobi, Renato
Publication Type Journal Items, Publication Status: Published
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Title Understanding chemical reactivity for homo- and heterobifunctional protein cross-linking agents
Author(s) Chen, Fan
Nielsen, Simone
Zenobi, Renato
Journal or Series Title Journal of mass spectrometry
Volume Number 48
Issue Number 7
Start Page 807
End Page 812
ISSN 1076-5174
Publisher Wiley
Publication Place Chichester
Publication Date 2013-07
Keyword(s) Chemical cross-linking
MALDI mass spectrometry
Amine-to-sulfhydryl cross-links
N-hydroxysuccinimide (NHS) esters
Abstract 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.
DOI 10.1002/jms.3224
Additional Notes Article first published online 18 June 2013, Manuscript revised 23 April 2013, Manuscript accepted 23 April 2013, Manuscript received 22 November 2012
Document Type Article
Publication Status Published
Language English
Assigned Organisational Unit(s) 03430
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NEBIS System Number 001410098
Source Database ID WOS-000321336000007
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  author = "Chen, Fan and Nielsen, Simone and Zenobi, Renato",
  title = "{U}nderstanding chemical reactivity for homo- and heterobifunctional protein cross-linking agents",
  journal = "Journal of mass spectrometry",
  year = 2013,
  volume = "48",
  number = "7",
  pages = "807--812",
  month = jul,

E-Citations record created: Mon, 12 Aug 2013, 07:26:31 CET