Use of de novo sequencing for proteins identification

Skvortsov V.S.1 , Mikurova A.V.1, Rybina A.V.1

1. Institute of Biomedical Chemistry, Moscow, Russia
DOI: 10.18097/PBMC20176304341      PubMed Id: 28862606
Year: 2017  Volume: 63  Issue: 4  Pages: 341-350
Three de novo sequencing programs (Novor, PEAKS and PepNovo+) have been used for identification of 48 individual human proteins constituting the Universal Proteomics Standard Set 2 (UPS2) (“Sigma-Aldrich”, USA). Experimental data have been obtained by tandem mass spectrometry. The MS/MS was performed using pure UPS2 and UPS2 mixtures with E. coli extract and human plasma samples. Protein detection was based on identification of at least two peptides of 9 residues in length or one peptide containing at least 13 residues. Using these criteria 13 (Novor), 20 (PEAKS) and 11 (PepNovo+) proteins were detected in pure UPS2 sample. Protein identifications in mixed samples were comparable or worse. Better results (by ~20%) were obtained using prediction included high quality identified fragment (TAG) containing at least 7 residues and unidentified additional masses at N- and C-termini (PepNovo+). The latter approach confidently recognized mass-spectrometric artefacts (and probably PTM). Atypical mass changes missed in UNIMOD DB were found (PepNovo+) to be statistically significant at the C-terminus (+23.02, +26.04 and +27.03). Using peptides containing these modifications and milder detection threshold 41 of 48 UPS2 proteins were identified.
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Keywords: shotgun proteomics, de novo sequencing, mass spectrometry, data processing

Skvortsov, V. S., Mikurova, A. V., Rybina, A. V. (2017). Use of de novo sequencing for proteins identification. Biomeditsinskaya khimiya, 63(4), 341-350.
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