DTASelect and Contrast:  Tools for Assembling and Comparing Protein Identifications from Shotgun Proteomics

Tabb, David L.; McDonald, W. Hayes; Jr, Yates

doi:10.1021/pr015504q

Cited by 1,356 publications

(1,197 citation statements)

References 17 publications

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“…In addition, the protein sequence database was concatenated to a reversed version of the database in order to define Xcorr and DeltaCN values for an appropriate false positive rate of protein identification. DTASelect [25] was used to apply these identification criteria in a manner similar to that employed by Peng et al [26]. No cleavage bias was assumed when searching MS/MS spectra against the S. mansoni sequence database.…”

Section: Mudpit Analysis Of Espmentioning

confidence: 99%

Proteomic analysis of Schistosoma mansoni egg secretions

Cass

Johnson

Califf

et al. 2007

Molecular and Biochemical Parasitology

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Schistosomiasis remains a largely neglected, global health problem. The morbid pathology of the disease stems from the host's inflammatory response to parasite eggs trapped in host tissues. Long term host/parasite survival is dependent upon the successful modulation of the acute pathological response, which is induced by egg antigens. In this study, using Multidimensional Protein Identification Technology, we identified the Schistosoma mansoni egg secretome consisting of 188 proteins. Notably we identified proteins involved in redox balance, molecular chaperoning and protein folding, development and signaling, scavenging and metabolic pathways, immune response modulation, and 32 novel, previously uncharacterized schistosome proteins. We localized a subset of previously-characterized schistosome proteins identified in egg secretions in this study, to the surface of live S. mansoni eggs using the circumoval precipitin reaction. The identification of proteins actively secreted by live schistosome eggs provides important new information for understanding immune modulation and the pathology of schistosomiasis.

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Section: Mudpit Analysis Of Espmentioning

confidence: 99%

Proteomic analysis of Schistosoma mansoni egg secretions

Cass

Johnson

Califf

et al. 2007

Molecular and Biochemical Parasitology

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194

196

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“…Next, the candidate proteins are sorted based on the cumulative number of matching spectra obtained°at°the°organelle°level°(Figure°2,°Table°1°and Table°2),°computational°procedures°are°carried°out°on data that have been formatted and parsed into an SQL-style relational database. Although this database was°developed°for°local°use,°the°reader°is°directed°to°the many helpful stand-alone informatics software tools presently available for managing, assessing, and filtering large-scale MudPIT-type proteomic datasets that are freely available to academics from the Yates and Aebersold°research°laboratories°(e.g.,°DTASelect° [34]; http://fields.scripps.edu/°and°ProteinProphet° [35]; www.systemsbiology.org/).…”

Section: Database Searching and Statistical Validationmentioning

confidence: 99%

Multidimensional protein identification technology (MudPIT): Technical overview of a profiling method optimized for the comprehensive proteomic investigation of normal and diseased heart tissue

Kislinger

Gramolini

MacLennan

et al. 2005

J. Am. Soc. Mass Spectrom.

133

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An optimized analytical expression profiling strategy based on gel-free multidimensional protein identification technology (MudPIT) is reported for the systematic investigation of biochemical (mal)-adaptations associated with healthy and diseased heart tissue. Enhanced shotgun proteomic detection coverage and improved biological inference is achieved by pre-fractionation of excised mouse cardiac muscle into subcellular components, with each organellar fraction investigated exhaustively using multiple repeat MudPIT analyses. Functional-enrichment, high-confidence identification, and relative quantification of hundreds of organelle-and tissue-specific proteins are achieved readily, including detection of low abundance transcriptional regulators, signaling factors, and proteins linked to cardiac disease. Important technical issues relating to data validation, including minimization of artifacts stemming from biased under-sampling and spurious false discovery, together with suggestions for further fine-tuning of sample preparation, are discussed. A framework for follow-up bioinformatic examination, pattern recognition, and data mining is also presented in the context of a stringent application of MudPIT for probing fundamental aspects of heart muscle physiology as well as the discovery of perturbations associated with heart failure. (J Am Soc Mass Spectrom 2005, 16, 1207-1220

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“…In evaluating spectra for inclusion, two different sets of criteria were used in the DTASelect and Contrast algorithms. 18 The "low" set used the settings −1 0.3 −2 0.3 −3 0.3 −Smn 5. This setting imposes a moderate cutoff normalized XCorr of 0.3 for peptide inclusion, limiting the listed peptides to sequences containing at least five residues and accepting the default of two peptides per protein for listed proteins.…”

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confidence: 99%

Influence of Basic Residue Content on Fragment Ion Peak Intensities in Low-Energy Collision-Induced Dissociation Spectra of Peptides

et al. 2004

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The primary utility of trypsin digestion in proteomics is that it cleaves proteins at predictable locations, but it is also notable for yielding peptides that terminate in basic arginine and lysine residues. Tryptic peptides fragment in ion trap tandem mass spectrometry to produce prominent Cterminal y series ions. Alternative proteolytic digests may produce peptides that do not follow these rules. In this study, we examine 2568 peptides generated through proteinase K digestion, a technique that produces a greater diversity of basic residue content in peptides. We show that the position of basic residues within peptides influences the peak intensities of b and y series ions; a basic residue near the N-terminus of a peptide can lead to prominent b series peaks rather than the intense y series peaks associated with tryptic peptides. The effects of presence and position for arginine, lysine, and histidine are explored separately and in combination. Arg shows the most dominant effects followed by His and then by Lys. Fragment ions containing basic residues produce more intense peaks than those without basic residues. Doubly charged precursor ions have generally been modeled as producing only singly charged fragment ions, but fragment ions that contain two basic residues may accept both protons during fragmentation. By characterizing the influence of basic residues on gasphase fragmentation of peptides, this research makes possible more accurate fragmentation models for peptide identification algorithms.Trypsin digestion is a standard step in many proteomic experiments. Proteins digested by this enzyme are predominantly cleaved C-terminal to arginine and lysine residues, yielding predictable collections of peptides.1 Tryptic peptides are particularly favored for tandem mass spectrometry because they yield spectra with prominent y series ions. Database search algorithms such as SEQUEST 2 have been tuned to work best with tryptic peptides. Alternative digestion techniques can be useful to analyze proteins that do not have many suitable cleavage sites for trypsin (such as membrane proteins) or to increase the diversity of peptides covering a protein region of interest. The cleavage specificities of other digestion enzymes may yield peptides that fragment differently than those produced by trypsin.One technique showing promise for proteomics is proteinase K digestion. 3 Proteinase K's specificity is more variable than trypsin, favoring amino acids with aromatic side chains as cleavage sites but targeting other residues as well. 4 The digestion time required is less than that of trypsin. Because of proteinase K's broader specificity, the diversity of peptides yielded by these digests is higher than in tryptic digests.The most common type of fragmentation used for proteomics is low-energy collision-induced dissociation (CID). In this technique, peptide ions of a particular mass-to-charge ratio are isolated and collided with noble gas atoms. Collisions add energy to the peptide ions and they fragment. The cleavage o...

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DTASelect and Contrast: Tools for Assembling and Comparing Protein Identifications from Shotgun Proteomics

Cited by 1,356 publications

References 17 publications

Proteomic analysis of Schistosoma mansoni egg secretions

Proteomic analysis of Schistosoma mansoni egg secretions

Multidimensional protein identification technology (MudPIT): Technical overview of a profiling method optimized for the comprehensive proteomic investigation of normal and diseased heart tissue

Influence of Basic Residue Content on Fragment Ion Peak Intensities in Low-Energy Collision-Induced Dissociation Spectra of Peptides

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