An EThcD-Based Method for Discrimination of Leucine and Isoleucine Residues in Tryptic Peptides

  • Sergey S. Zhokhov
  • Sergey V. Kovalyov
  • Tatiana Yu. Samgina
  • Albert T. Lebedev
Research Article
  • 303 Downloads

Abstract

An EThcD-based approach for the reliable discrimination of isomeric leucine and isoleucine residues in peptide de novo sequencing procedure has been proposed. A multistage fragmentation of peptide ions was performed with Orbitrap Elite mass spectrometer in electrospray ionization mode. At the first stage, z-ions were produced by ETD or ETcaD fragmentation of doubly or triply charged peptide precursor ions. These primary ions were further fragmented by HCD with broad-band ion isolation, and the resulting w-ions showed different mass for leucine and isoleucine residues. The procedure did not require manual isolation of specific z-ions prior to HCD stage. Forty-three tryptic peptides (3 to 27 residues) obtained by trypsinolysis of human serum albumin (HSA) and gp188 protein were analyzed. To demonstrate a proper solution for radical site migration problem, three non-tryptic peptides were also analyzed. A total of 93 leucine and isoleucine residues were considered and 83 of them were correctly identified. The developed approach can be a reasonable substitution for additional Edman degradation procedure, which is still used in peptide sequencing for leucine and isoleucine discrimination.

Graphical Abstract

Keywords

Leucine/isoleucine differentiation Orbitrap EThcD Tryptic peptides Peptide sequencing 

Supplementary material

13361_2017_1674_MOESM1_ESM.doc (1.7 mb)
ESM 1(DOC 1.67 MB)

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Copyright information

© American Society for Mass Spectrometry 2017

Authors and Affiliations

  • Sergey S. Zhokhov
    • 1
  • Sergey V. Kovalyov
    • 1
  • Tatiana Yu. Samgina
    • 1
  • Albert T. Lebedev
    • 1
  1. 1.Department of ChemistryM.V. Lomonosov Moscow State UniversityMoscowRussia

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