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Monitoring Functional Posttranslational Modifications Using a Data-Driven Proteome Informatic Pipeline
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DOI：10.1002/pmic.202400238.png)
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Posttranslational modifications (PTMs) are of significant interest in molecular biomedicine due to their crucial role in signal transduction across various cellular and organismal processes. Characterizing PTMs, distinguishing between functional and inert modifications, quantifying their occupancies, and understanding PTM crosstalk are challenging tasks in any biosystem. Studying each PTM often requires a specific, labor-intensive experimental design. Here, we present a PTM-centric proteome informatic pipeline for predicting relevant PTMs in mass spectrometry-based proteomics data without prior information. Once predicted, these in silico identified PTMs can be incorporated into a refined database search and compared to measured data. As a practical application, we demonstrate how this pipeline can be used to study glycoproteomics in oral squamous cell carcinoma based on the proteome profile of primary tumors. Subsequently, we experimentally identified cellular proteins that are differentially expressed in cells treated with multikinase inhibitors dasatinib and staurosporine using mass spectrometry-based proteomics. Computational enrichment analysis was then employed to determine the potential PTMs of differentially expressed proteins induced by both drugs. Finally, we conducted an additional round of database search with the predicted PTMs. Our pipeline successfully analyzed the enriched PTMs, and detected proteins not identified in the initial search. Our findings support the effectiveness of PTM-centric searching of MS data in proteomics based on computational enrichment analysis, and we propose integrating this approach into future proteomics search engines.
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PEPTIDE IDENTIFICATION
ENRICHMENT
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