Mass spectrometric de novo sequencing of natural non‐tryptic peptides: comparing peculiarities of collision‐induced dissociation (CID) and high energy collision dissociation (HCD)

peer-reviewed · Rapid Communications in Mass Spectrometry · 2014

peer-reviewed · Rapid Communications in Mass Spectrometry · 2014. Tatiana Yu. Samgina et al. Rationale Mass spectrometry has shown itself as the most efficient tool for the sequencing of peptides…
Date 2014-12-15
Type peer-reviewed
Venue Rapid Communications in Mass Spectrometry
Publisher Wiley
Contribution adjacent
DOI 10.1002/rcm.7049
Citations (OpenAlex) 25
Venue 2-year citedness 1.80

Abstract

Rationale Mass spectrometry has shown itself as the most efficient tool for the sequencing of peptides. However, de novo sequencing of novel natural peptides is significantly more challenging in comparison with the same procedure applied for the tryptic peptides. To reach the goal in this case it is essential to select the most useful methods of triggering fragmentation and combine complementary techniques. Methods Comparison of low-energy collision-induced dissociation (CID) and higher energy collision-induced dissociation (HCD) modes for sequencing of the natural non-tryptic peptides with disulfide bonds and/or several proline residues in the backbone was achieved using an LTQ FT Ultra Fourier transform ion cyclotron resonance (FTICR) mass spectrometer (Thermo Fisher Scientific, Bremen, Germany) equipped with a 7 T magnet and an LTQ Orbitrap Velos ETD (Thermo Fisher Scientific, Bremen, Germany) instrument. Peptide fractions were obtained by high-performance liquid chromatography (HPLC) separation of frog skin secretion samples from ten species of Rana temporaria, caught in the Kolomna district of Moscow region (Russia). Results HCD makes the b/y series longer and more pronounced, thus increasing sequence coverage. Fragment ions due to cleavages at the C-termini of proline residues make the sequencing more reliable and may be used to detect missed cleavages in the case of tryptic peptides. Another HCD peculiarity involves formation of pronounced inner fragment ions (secondary y(n)b(m) ion series formed from the abundant primary y-ions). Differences in de novo sequencing of natural non-tryptic peptides with CID and HCD, involving thorough manual expert interpretation of spectra and two automatic sequencing algorithms, are discussed. Conclusions Although HCD provides better results, a combination of CID and HCD data may notably increase reliability of de novo sequencing. Several pairs of b2 /a2 -ions may be formed in HCD, complicating the spectra. Automatic de novo sequencing with the available programs remains less efficient than the manual one, independently of the collision energy.

Authors

  1. Tatiana Yu. Samgina · Lomonosov Moscow State University, MSU-BIT University
  2. Egor A. Vorontsov · Lomonosov Moscow State University
  3. Vladimir Gorshkov · Justus-Liebig-Universität Gießen, Lomonosov Moscow State University, University of Southern Denmark
  4. Konstantin A. Artemenko · Lomonosov Moscow State University, Uppsala University
  5. Roman A. Zubarev · Cornell University, Karolinska Institutet, University of Southern Denmark, Uppsala University
  6. Albert T. Lebedev · Lomonosov Moscow State University, MSU-BIT University

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