Dataset Information

Detecting diagnostic features in MS/MS spectra of post-translationally modified peptides.

ABSTRACT: Post-translational modifications are an area of great interest in mass spectrometry-based proteomics, with a surge in methods to detect them in recent years. However, post-translational modifications can introduce complexity into proteomics searches by fragmenting in unexpected ways, ultimately hindering the detection of modified peptides. To address these deficiencies, we present a fully automated method to find diagnostic spectral features for any modification. The features can be incorporated into proteomics search engines to improve modified peptide recovery and localization. We show the utility of this approach by interrogating fragmentation patterns for a cysteine-reactive chemoproteomic probe, RNA-crosslinked peptides, sialic acid-containing glycopeptides, and ADP-ribosylated peptides. We also analyze the interactions between a diagnostic ion's intensity and its statistical properties. This method has been incorporated into the open-search annotation tool PTM-Shepherd and the FragPipe computational platform.

SUBMITTER: Geiszler DJ

PROVIDER: S-EPMC10338467 | biostudies-literature | 2023 Jul

REPOSITORIES: biostudies-literature

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Detecting diagnostic features in MS/MS spectra of post-translationally modified peptides.

Geiszler Daniel J DJ Polasky Daniel A DA Yu Fengchao F Nesvizhskii Alexey I AI

Nature communications 20230712 1

Post-translational modifications are an area of great interest in mass spectrometry-based proteomics, with a surge in methods to detect them in recent years. However, post-translational modifications can introduce complexity into proteomics searches by fragmenting in unexpected ways, ultimately hindering the detection of modified peptides. To address these deficiencies, we present a fully automated method to find diagnostic spectral features for any modification. The features can be incorporated ...[more]

PMID: 37438360

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Project description:Peptides presented by MHC molecules on the cell surface, or the immunopeptidome, play an important role in the adaptive arm of the immune response. Antigen processing for MHC class I molecules is a ubiquitous pathway present in all nucleated cells which generates and presents peptides of both self and non-self-origin. Peptides with post-translational modifications represent one category of peptides presented by MHC class I molecules. However, owing to the complexity of self-peptides presented by cells, the diversity of peptides with post-translational modifications is not well-studied. In this study, we carried out MHC Class I immunopeptidomics analysis of Loucy T-cell leukemia and A375 malignant melanoma cell line to characterize the diversity of post-translational modifications of MHC class I-bound peptides. Using high resolution mass spectrometry, we identified 25,761 MHC-bound peptides across both cell lines using Bolt and Sequest search engines. The enrichment method was highly specific as ~ 90% of the peptides were of typical length (8-12 amino acids long) and the motifs were expected based on previously reported motifs for MHC I alleles. Among the MHC-bound peptides, we identified phosphorylation as a major post-translational modification followed by deamidation. We observed site-specific localization of these post-translational modifications, at position P4 for phosphorylated peptides and position P3 for deamidated peptides. We identified a smaller number of peptides with acetylated and methylated lysine, possibly due to very low stoichiometric levels of these PTMs compared to phosphorylation and deamidation. Using PEAKS de novo sequencing algorithm, we identified spliced peptides that accounted for ~ 5-7% of MHC-bound peptides that were otherwise similar in their features as normal MHC-bound peptides. We validated the identity of several post-translationally modified peptides and spliced peptides through mass spectrometric analysis of synthetic peptides. Our study confirms post-translationally modified peptides to be present at low stoichiometric levels along with unusual spliced peptides through unbiased identification using high resolution mass spectrometry.Supplementary informationThe online version contains supplementary material available at 10.1007/s42485-021-00066-x.

Dataset Information

Detecting diagnostic features in MS/MS spectra of post-translationally modified peptides.

Publications

Detecting diagnostic features in MS/MS spectra of post-translationally modified peptides.

Similar Datasets

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