A high-throughput de novo sequencing approach for shotgun proteomics using high-resolution tandem mass spectrometry

Abstract

BackgroundHigh-resolution tandem mass spectra can now be readily acquired with hybrid instruments, such as LTQ-Orbitrap and LTQ-FT, in high-throughput shotgun proteomics workflows. The improved spectral quality enables more accurate de novo sequencing for identification of post-translational modifications and amino acid polymorphisms.ResultsIn this study, a new de novo sequencing algorithm, called Vonode, has been developed specifically for analysis of such high-resolution tandem mass spectra. To fully exploit the high mass accuracy of these spectra, a unique scoring system is proposed to evaluate sequence tags based primarily on mass accuracy information of fragment ions. Consensus sequence tags were inferred for 11,422 spectra with an average peptide length of 5.5 residues from a total of 40,297 input spectra acquired in a 24-hour proteomics measurement of Rhodopseudomonas palustris. The accuracy of inferred consensus sequence tags was 84%. According to our comparison, the performance of Vonode was shown to be superior to the PepNovo v2.0 algorithm, in terms of the number of de novo sequenced spectra and the sequencing accuracy.ConclusionsHere, we improved de novo sequencing performance by developing a new algorithm specifically for high-resolution tandem mass spectral data. The Vonode algorithm is freely available for download at http://compbio.ornl.gov/Vonode.

JUMP: A Tag-based Database Search Tool for Peptide Identification with High Sensitivity and Accuracy

“…De novo methods, including Lutefisk (29), PEAKS (30), NovoHMM (31), PepNovo (32), pNovo (33), Vonovo (34), and UniNovo (35), identify peptide sequences directly from MS/MS spectra. These methods can be used to derive novel peptides and post-translational modifications without a database, which is useful, especially when the related genome is not sequenced.…”

De novo sequencing of proteins and peptides: algorithms, applications, perspectives

“…Подавляющее их большинство было разработано для установления первичной структуры пептидов по данным масс-спектрометрии "снизу вверх" (bottom-up). К тем из них, что оказались наиболее привлекательными для специалистов, могут быть отнесены Lutefisk [26,39], SHERENGA [40], PEAKS [28], PepNovo [29] и усовершенствованная его версия PepNovo+ [41][42][43], NovoHMM [44], Vonode [23], pNovo [45], UniNovo [46] и Novor [34].…”

“…При этом в рассмотрении остаются лишь те аминокислотные последовательности из базы, которые согласуются с найденными тегами. Данный подход был впервые предложен в 1994 г. М. Манном и М. Вилмом [12], а впоследствии были опубликованы десятки статей, в которых обсуждались эффективные способы генерации тегов и их использования в качестве фильтров (см., например, [13][14][15][16][17][18][19][20][21][22][23][24][25] и [8]). …”

De Novo Sequencing and Homology Searching

“…Lutefisk1900 then became a benchmark that allowed others to show the superiority of their own programs. Many de novo sequencing programs (33)(34)(35)(36)(37)(38)(39)(40)(41)(42)(43) have been developed since these early programs. Among these programs the better known were Sherenga (39), PEAKS (37) and PepNovo (36), which employ more sensible probabilistic scoring schemes and are generally faster than Lutefisk.…”