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Electron capture dissociation proceeds with a low degree of intramolecular migration of peptide amide hydrogens
Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Pharmacy, Department of Pharmaceutical Biosciences, MMS, Medical Mass Spectrometry.
Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Pharmacy, Department of Pharmaceutical Biosciences, MMS, Medical Mass Spectrometry.
2008 (English)In: Journal of the American Chemical Society, ISSN 0002-7863, E-ISSN 1520-5126, Vol. 130, no 4, 1341-1349 p.Article in journal (Refereed) Published
Abstract [en]

Hydrogen (H-1/H-2) exchange combined with mass spectrometry (HX-MS) has become a recognized method for the analysis of protein structural dynamics. Presently, the incorporated deuterons are typically localized by enzymatic cleavage of the labeled proteins and single residue resolution is normally only obtained for a few residues. Determination of site-specific deuterium levels by gas-phase fragmentation in tandem mass spectrometers would greatly increase the applicability of the HX-MS method. The biggest obstacle in achieving this goal is the intramolecular hydrogen migration (i.e., hydrogen scrambling) that occurs during vibrational excitation of gas-phase ions. Unlike traditional collisional ion activation, electron capture dissociation (ECD) is not associated with substantial vibrational excitation. We investigated the extent of intramolecular backbone amide hydrogen (H-1/H-2) migration upon ECD using peptides with a unique selective deuterium incorporation. Our results show that only limited amide hydrogen migration occurs upon ECD, provided that vibrational excitation prior to the electron capture event is minimized. Peptide ions that are excessively vibrationally excited in the electrospray ion source by, e.g., high declustering potentials or during precursor ion selection (via side band excitation) in the external linear quadrupole ion trap undergo nearly complete hydrogen (H-1/H-2) scrambling. Similarly, collision-induced dissociation (CID) in the external linear quadrupole ion trap results in complete or extensive hydrogen (H-1/H-2) scrambling. This precludes the use of CID as a method to obtain site-specific information from proteins that are labeled in solution-phase H-1/H-2 exchange experiments. In contrast, the deuteration levels of the c- and z-fragment ions generated from ECD closely mimic the known solution deuteration pattern of the selectively labeled peptides. This excellent correlation between the results obtained from gas phase and solution suggests that ECD holds great promise as a general method to obtain single residue resolution in proteins from solution H-1/H-2 exchange experiments.

Place, publisher, year, edition, pages
2008. Vol. 130, no 4, 1341-1349 p.
National Category
Biological Sciences
Identifiers
URN: urn:nbn:se:uu:diva-149889DOI: 10.1021/ja076448iISI: 000252634600045PubMedID: 18171065OAI: oai:DiVA.org:uu-149889DiVA: diva2:405882
Available from: 2011-03-24 Created: 2011-03-24 Last updated: 2017-12-11Bibliographically approved

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