Both authors contributed equally to this work.
Collision-induced dissociative chemical cross-linking reagent for protein structure characterization: applied Edman chemistry in the gas phase
Version of Record online: 30 NOV 2009
Copyright © 2009 John Wiley & Sons, Ltd.
Journal of Mass Spectrometry
Volume 45, Issue 2, pages 178–189, February 2010
How to Cite
Dreiocker, F., Müller, M. Q., Sinz, A. and Schäfer, M. (2010), Collision-induced dissociative chemical cross-linking reagent for protein structure characterization: applied Edman chemistry in the gas phase. J. Mass Spectrom., 45: 178–189. doi: 10.1002/jms.1702
- Issue online: 9 FEB 2010
- Version of Record online: 30 NOV 2009
- Manuscript Accepted: 1 NOV 2009
- Manuscript Received: 25 AUG 2009
- German Research Foundation (DFG)
- chemical cross-linking;
- Edman chemistry;
- collision-induced dissociation;
- protein structure analysis
Chemical cross-linking combined with a subsequent enzymatic digestion and mass spectrometric analysis of the created cross-linked products presents an alternative approach to assess low-resolution protein structures and to gain insight into protein interfaces. In this contribution, we report the design of an innovative cross-linker based on Edman degradation chemistry, which leads to the formation of indicative mass shifted fragment ions and constant neutral losses (CNLs) in electrospray ionization (ESI)-tandem-mass spectrometry (MS/MS) product ion mass spectra, allowing an unambiguous identification of cross-linked peptides. Moreover, the characteristic neutral loss reactions facilitate automated analysis by multiple reaction monitoring suited for high throughput studies with good sensitivity and selectivity. The functioning of the novel cross-linker relies on the presence of a highly nucleophilic sulfur in a thiourea moiety, safeguarding for effective intramolecular attack leading to predictive and preferred cleavage of a glycyl-prolyl amide bond. Our innovative analytical concept and the versatile applicability of the collision-induced dissociative chemical cross-linking reagent are exemplified for substance P, luteinizing hormone releasing hormone LHRH and lysozyme. The novel cross-linker is expected to have a broad range of applications for probing protein tertiary structures and for investigating protein–protein interactions. Copyright © 2009 John Wiley & Sons, Ltd.