Discrimination of Leucine and Isoleucine in Peptides Sequencing with Orbitrap Fusion Mass Spectrometer
peer-reviewed · Analytical Chemistry · 2014
| Date | 2014-07-15 |
| Type | peer-reviewed |
| Venue | Analytical Chemistry |
| Publisher | American Chemical Society (ACS) |
| Contribution | adjacent |
| DOI | 10.1021/ac501200h |
| Citations (OpenAlex) | 92 |
| Venue 2-year citedness | 7.10 |
Abstract
An efficient approach to easy and reliable differentiation between isomeric leucine and isoleucine in peptide sequencing utilizes multistage electron transfer dissociation and higher energy collision activated dissociation in the Orbitrap Fusion mass spectrometer. The MS(3) method involves production and isolation of primary odd-electron z(•) ions, followed by radical site initiation of their fragmentation with formation of w-ions, characteristic of the isomeric amino acid residues. Six natural nontryptic peptides isolated from the secretion of frog Rana ridibunda were studied. Their lengths were in the range between 15 and 37 amino acids and the number of targeted isomeric (Leu/Ile) residues varied between 1 and 7. The experiments were successful in all 22 cases of Leu/Ile residues, leaving no doubts in identification. The method is extremely selective as the targeted w-ions appear to be the most intense in the spectra. The proposed approach may be incorporated into shotgun proteomics algorithms and allows for the development of an exclusively mass spectrometric method for automated complete de novo sequencing of various peptides and proteins.
Methods and tools
- MS3 w-ion Leu/Ile discrimination: Separates the isomeric residues leucine and isoleucine during de novo sequencing by instrument rather than by inference: multistage ETD isolates the primary odd-electron z-dot ions and HCD then drives radical-site fragmentation into w-ions, whose masses differ between the two isomers and which come out as the most intense peaks in the spectrum. Demonstrated on six natural non-tryptic peptides of 15 to 37 residues from Rana ridibunda frog skin secretion, resolving all 22 Leu/Ile positions.
Cites (1)
Cited by (5)
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