Pepitome: Evaluating Improved Spectral Library Search for Identification Complementarity and Quality Assessment

Dasari, Surendra; Chambers, Matthew; Martinez, Misti A.; Carpenter, Kristin L.; Ham, Amy Joan L.; Vega‐Montoto, Lorenzo; Tabb, David L.

doi:10.1021/pr200874e

Cited by 60 publications

(70 citation statements)

References 34 publications

(89 reference statements)

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“…The mzml files were searched using MyriMatch version 2.1.132 (33), Pepitome version 1.0.42 (34), Comet version 2014.01.1 (35), and MS-GFϩ version 9517 (36) against the human Refseq version 54 database (Sep 25, 2012; 34,590 entries). A semitryptic search was employed with a maximum of four missed cleavages allowed.…”

Section: Methodsmentioning

confidence: 99%

Assembly Dynamics and Stoichiometry of the Apoptosis Signal-regulating Kinase (ASK) Signalosome in Response to Electrophile Stress

Federspiel

Codreanu

Palubinsky

et al. 2016

Molecular & Cellular Proteomics

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Apoptosis signal-regulating kinase 1 (ASK1) is a key sensor kinase in the mitogen-activated protein kinase pathway that transduces cellular responses to oxidants and electrophiles. ASK1 is regulated by a large, dynamic multiprotein signalosome complex, potentially including over 90 reported ASK1-interacting proteins. We employed both shotgun and targeted mass spectrometry assays to catalogue the ASK1 protein-protein interactions in HEK-293 cells treated with the prototypical lipid electrophile 4-hydroxy-2-nonenal (HNE). Using both epitope-tagged overexpression and endogenous expression cell systems, we verified most of the previously reported ASK1 proteinprotein interactions and identified 14 proteins that exhibited dynamic shifts in association with ASK1 in response to HNE stress. We used precise stable isotope dilution assays to quantify protein stoichiometry in the ASK signalosome complex and identified ASK2 at a 1:1 stoichiometric ratio with ASK1 and 14 -3-3 proteins (YWHAQ, YWHAB, YWHAH, and YWHAE) collectively at a 0.5:1 ratio with ASK1 as the main components. Several other proteins, including ASK3, PARK7, PRDX1, and USP9X were detected with stoichiometries of 0

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Section: Methodsmentioning

confidence: 99%

Assembly Dynamics and Stoichiometry of the Apoptosis Signal-regulating Kinase (ASK) Signalosome in Response to Electrophile Stress

Federspiel

Codreanu

Palubinsky

et al. 2016

Molecular & Cellular Proteomics

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“…Some other programs, such as SpectraST (21) and Pepitome (22), utilize a spectral library composed of experimentally identified and validated MS/MS spectra. These methods use a variety of scoring algorithms to rank potential peptide spectrum matches (PSMs) and select the top hit as a putative PSM.…”

mentioning

confidence: 99%

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

Wang

et al. 2014

Molecular & Cellular Proteomics

129

124

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Database search programs are essential tools for identifying peptides via mass spectrometry (MS) in shotgun proteomics. Simultaneously achieving high sensitivity and high specificity during a database search is crucial for improving proteome coverage. Here we present JUMP, a new hybrid database search program that generates amino acid tags and ranks peptide spectrum matches (PSMs) by an integrated score from the tags and pattern matching. In a typical run of liquid chromatography coupled with high-resolution tandem MS, more than 95% of MS/MS spectra can generate at least one tag, whereas the remaining spectra are usually too poor to derive genuine PSMs. To enhance search sensitivity, the JUMP program enables the use of tags as short as one amino acid. Using a target-decoy strategy, we compared JUMP with other programs (e.g. SEQUEST, Mascot, PEAKS DB, and InsPecT) in the analysis of multiple datasets and found that JUMP outperformed these preexisting programs. JUMP also permitted the analysis of multiple co-fragmented peptides from "mixture spectra" to further increase PSMs. In addition, JUMP-derived tags allowed partial de novo sequencing and facilitated the unambiguous assignment of modified residues. In summary, JUMP is an effective database search algorithm complementary to current search programs. Molecular & Cellular Proteomics 13: 10.1074/mcp.O114.039586, 3663-3673, 2014.Peptide identification by tandem mass spectra is a critical step in mass spectrometry (MS)-based 1 proteomics (1). Numerous computational algorithms and software tools have been developed for this purpose (2-6). These algorithms can be classified into three categories: (i) pattern-based database search, (ii) de novo sequencing, and (iii) hybrid search that combines database search and de novo sequencing. With the continuous development of high-performance liquid chromatography and high-resolution mass spectrometers, it is now possible to analyze almost all protein components in mammalian cells (7). In contrast to rapid data collection, it remains a challenge to extract accurate information from the raw data to identify peptides with low false positive rates (specificity) and minimal false negatives (sensitivity) (8).Database search methods usually assign peptide sequences by comparing MS/MS spectra to theoretical peptide spectra predicted from a protein database, as exemplified in SEQUEST (9), Mascot (10), OMSSA (11), X!Tandem (12), Spectrum Mill (13), ProteinProspector (14), MyriMatch (15), Crux (16), MS-GFDB (17), Andromeda (18), BaMS 2 (19), and Morpheus (20). Some other programs, such as SpectraST (21) and Pepitome (22), utilize a spectral library composed of experimentally identified and validated MS/MS spectra. These methods use a variety of scoring algorithms to rank potential peptide spectrum matches (PSMs) and select the top hit as a putative PSM. However, not all PSMs are correctly assigned. For example, false peptides may be assigned to MS/MS spectra with numerous noisy peaks and poor fragmentation patterns. If the samples conta...

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“…Shotgun proteomics has a number of advantages: not using 2-DE; analyses of basic proteins; analysis of high molecular weight proteins (>120 kDa); and detection of low abundance proteins [28]. Dasari et al [29] developed Pepitome, a new spectral library search tool, for the identification of peptides by comparing experimental MS/MS scans to those in spectral libraries. This tool makes the automation of quality analysis and quality control for shotgun proteomics data possible [29].…”

Section: Discovery and Verification Approachesmentioning

confidence: 99%

“…Dasari et al [29] developed Pepitome, a new spectral library search tool, for the identification of peptides by comparing experimental MS/MS scans to those in spectral libraries. This tool makes the automation of quality analysis and quality control for shotgun proteomics data possible [29]. Despite the advances such as high-throughput, high sensitivity and precision, these methods have some disadvantages.…”

Section: Discovery and Verification Approachesmentioning

confidence: 99%

Progress and Clinical Applications in Proteomics

Karasawa¹

2013

J Data Mining in Genom Proteomics

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Pepitome: Evaluating Improved Spectral Library Search for Identification Complementarity and Quality Assessment

Cited by 60 publications

References 34 publications

Assembly Dynamics and Stoichiometry of the Apoptosis Signal-regulating Kinase (ASK) Signalosome in Response to Electrophile Stress

Assembly Dynamics and Stoichiometry of the Apoptosis Signal-regulating Kinase (ASK) Signalosome in Response to Electrophile Stress

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

Progress and Clinical Applications in Proteomics

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