pNovo: De novo Peptide Sequencing and Identification Using HCD Spectra

Abstract

De novo peptide sequencing has improved remarkably in the past decade as a result of better instruments and computational algorithms. However, de novo sequencing can correctly interpret only ∼30% of high-and medium-quality spectra generated by collision-induced dissociation (CID), which is much less than database search. This is mainly due to incomplete fragmentation and overlap of different ion series in CID spectra. In this study, we show that higher-energy collisional dissociation (HCD) is of great help to de novo sequencing because it produces high mass accuracy tandem mass spectrometry (MS/MS) spectra without the low-mass cutoff associated with CID in ion trap instruments. Besides, abundant internal and immonium ions in the HCD spectra can help differentiate similar peptide sequences. Taking advantage of these characteristics, we developed an algorithm called pNovo for efficient de novo sequencing of peptides from HCD spectra. pNovo gave correct identifications to 80% or more of the HCD spectra identified by database search. The number of correct full-length peptides sequenced by pNovo is comparable with that obtained by database search. A distinct advantage of de novo sequencing is that deamidated peptides and peptides with amino acid mutations can be identified efficiently without extra cost in computation. In summary, implementation of the HCD characteristics makes pNovo an excellent tool for de novo peptide sequencing from HCD spectra.

Database independent proteomics analysis of the ostrich and human proteome

“…To reduce misassignments, approaches have been suggested based on improved mass accuracy or precision (14,15), combining complementary fragmentation data (14,16,17) or the simplification of fragmentation spectra by using chemical derivatization to manipulate peptide behavior (18). Recently, we introduced a biochemical approach to simplify fragment spectra based on the protease Lys-N (19), which has enzymatic cleavage specificity for the N-terminal side of lysine residues.…”

Rapid development of proteomics in China: from the perspective of the Human Liver Proteome Project and technology development

“…He Simin's and Dong Mengqiu's lab pioneered this direction in the field. They developed the search engine pNovo+ for de novo peptide sequencing with a greater than 70% success rate using HCD and ETD alone or complementary HCD and ETD tandem mass spectra [69][70][71]. More interestingly, they used pNovo+ to help identify a member of the serine protease inhibitor (Serpin) family, As_SRP-1, which is secreted from spermatids during spermiogenesis in the nematode Ascaris suum.…”

UniNovo : A Universal Tool for de Novo Peptide Sequencing

“…5.3, online) (Ma et al, 2003) and pNovo (ver. 1.1) (Chi et al, 2010) using the datasets in Table 1. For each tool, we generated N de novo reconstructions per each spectrum for N ¼ 1, 5 and 20.…”

“…For ETD2 and ETDL4 datasets, UniNovo . Per each spectrum, N top scoring reconstructions were generated by UniNovo, PepNovoþ (Frank, 2009;Frank and Pevzner, 2005), PEAKS (Ma et al, 2003), pNovo (Chi et al, 2010) and MS-GFDBScore. MS-GFDBScore provides UniNovo with MS-GFDB's scoring function.…”