pFind 2.0: a software package for peptide and protein identification via tandem mass spectrometry

Wang, Le-Heng; Li, Dequan; Fu, Yan; Wang, Haipeng; Zhang, Jingfen; Yuan, Zuo‐Fei; Sun, Rui-Xiang; Zeng, Rong; He, Shan; Gao, Wen

doi:10.1002/rcm.3173

Cited by 202 publications

(180 citation statements)

References 30 publications

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“…In all of the experiments, the most widely used software for peptide identification, Mascot (version 2.2) (7), was used to identify the spectra. Two other search engines, SEQUEST (version 2.7) (6) and pFind (version 2.6) (8,26,27), were additionally used to demonstrate the linearity of the ␥ k ͑x͒ function (Fig. 1).…”

Section: Resultsmentioning

confidence: 99%

Transferred Subgroup False Discovery Rate for Rare Post-translational Modifications Detected by Mass Spectrometry

Qian

2014

Molecular & Cellular Proteomics

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122

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In shotgun proteomics, high-throughput mass spectrometry experiments and the subsequent data analysis produce thousands to millions of hypothetical peptide identifications. The common way to estimate the false discovery rate (FDR) of peptide identifications is the target-decoy database search strategy, which is efficient and accurate for large datasets. However, the legitimacy of the target-decoy strategy for protein-modification-centric studies has rarely been rigorously validated. It is often the case that a global FDR is estimated for all peptide identifications including both modified and unmodified peptides, but that only a subgroup of identifications with a certain type of modification is focused on. As revealed recently, the subgroup FDR of modified peptide identifications can differ dramatically from the global FDR at the same score threshold, and thus the former, when it is of interest, should be separately estimated. However, rare modifications often result in a very small number of modified peptide identifications, which makes the direct separate FDR estimation inaccurate because of the inadequate sample size. This paper presents a method called the transferred FDR for accurately estimating the FDR of an arbitrary number of modified peptide identifications. Through flexible use of the empirical data from a targetdecoy database search, a theoretical relationship between the subgroup FDR and the global FDR is made computable. Through this relationship, the subgroup FDR can be predicted from the global FDR, allowing one to avoid an inaccurate direct estimation from a limited amount of data. The effectiveness of the method is demonstrated with both simulated and real mass spectra. Molecular & Cellular

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Section: Resultsmentioning

confidence: 99%

Transferred Subgroup False Discovery Rate for Rare Post-translational Modifications Detected by Mass Spectrometry

Qian

2014

Molecular & Cellular Proteomics

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122

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“…All mass spectra were acquired in the mass range of 200 to 2800 m/z at a scan speed of 8000 m/z per second. MS/MS spectra were searched using Mascot 2.3 and pFind Studio 2.8 software [44]. All tandem mass spectra were searched against an in-house database that contains all the protein sequences generated from A. bicolor transcriptomic analysis, and the peptides/proteins that were downloaded from UniProtKB.…”

Section: Proteomics Analysismentioning

confidence: 99%

Unique diversity of the venom peptides from the scorpion Androctonus bicolor revealed by transcriptomic and proteomic analysis

Zhang

Shi

Zeng

et al. 2015

Journal of Proteomics

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“…All of the raw files were converted to *.mgf files using pXtract (part of the pFind program 20 ). The intensity values of the peptide a, b, and y fragment ion pairs were extracted from the mgf files using java scripts built in-house to calculate the H/ L ratio of the peptide.…”

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confidence: 99%

Mass Defect-Based Pseudo-Isobaric Dimethyl Labeling for Proteome Quantification

Zhou

Shan

et al. 2013

Anal. Chem.

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Discovering differentially expressed proteins in various biological samples requires proteome quantification methods with accuracy, precision, and wide dynamic range. This study describes a mass defect-based pseudo-isobaric dimethyl labeling (pIDL) method based on the subtle mass defect differences between 12 C/ 13 C and 1 H/ 2 H. Lys-C protein digests were labeled with CD 2 O/ 13 CD 2 O and reduced with NaCNBD 3 /NaCNBH 3 as heavy and light isotopologues, respectively. The fragment ion pairs with mass differences of 5.84 mDa were resolved by high-resolution tandem mass spectrometry (MS/MS) and used for quantification. The pIDL method described here resulted in highly accurate and precise quantification results with approximately 100-fold dynamic range. Furthermore, the pIDL method was extended to 4-plex proteome quantification and applied to the quantitative analysis of proteomes from Hca-P and Hca-F, two mouse hepatocarcinoma ascites syngeneic cell lines with low and high lymph node metastasis rates.M ethods of stable-isotope incorporation with mass spectrometry (MS)-based proteome quantification have advanced rapidly in the past decade. Peptide samples can be differentially tagged with heavy or light isotopes by metabolic labeling 1,2 or chemical labeling. 3,4 The mass differences can be distinguished at either the MS or tandem mass spectrometry (MS/MS) level. Dimethyl labeling, 3 a chemical labeling method, is widely used for proteome quantification at the MS level. Several advantages of this method include quick reaction, high labeling efficiency, low cost, and applicability to different types of samples including tissues, cells, and body fluids. 5 Isobaric tags for relative and absolute quantitation (iTRAQ), 4 an MS/MS-based method, allows the proteome quantification of up to eight samples simultaneously and provides more precise quantification results than the MS level quantification method. 6,7 Although these strategies have been widely used for proteome quantification, their accuracy and dynamic range are limited by the signal-tonoise ratio and the increased MS spectral complexity leads to fewer quantified proteins for MS level-based quantification approaches. 8 In addition, ratio distortion caused by precursor interference is common for MS/MS level-based quantification methods. 9,10 To solve these problems, several innovative methods have been recently developed. Isobaric peptide termini labeling (IPTL) 11−13 is an elegant solution in which the amino groups of the N-termini and C-termini of Lys-C protein digests were crosswise labeled with heavy/light isotope reagents according to the slightly different chemical properties of α-and ε-NH 2 . Fragment ion pairs specific to the labeled peptides were used for peptide/protein quantification. Although IPTL improved the quantification accuracy compared to other reported MS/ MS level methods, the side reactions that are inherent in the multistep labeling can adversely affect quantification accuracy and dynamic range. 12 NeuCode SILAC 14 is another st...

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pFind 2.0: a software package for peptide and protein identification via tandem mass spectrometry

Cited by 202 publications

References 30 publications

Transferred Subgroup False Discovery Rate for Rare Post-translational Modifications Detected by Mass Spectrometry

Transferred Subgroup False Discovery Rate for Rare Post-translational Modifications Detected by Mass Spectrometry

Unique diversity of the venom peptides from the scorpion Androctonus bicolor revealed by transcriptomic and proteomic analysis

Mass Defect-Based Pseudo-Isobaric Dimethyl Labeling for Proteome Quantification

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