SeMoP: a new computational strategy for the unrestricted search for modified peptides using LC-MS/MS data

Area of Science:

• Proteomics
• Computational Biology
• Mass Spectrometry

Background:

• Identifying peptide modifications is crucial for understanding protein function and biological processes.
• Shotgun proteomics often faces challenges in comprehensively discovering and verifying diverse peptide modifications.
• Existing methods may have limitations in sensitivity and scope for unrestricted modification discovery.

Purpose of the Study:

• To present a novel computational approach, Search for Modified Peptides (SeMoP), for unrestricted discovery and verification of peptide modifications.
• To demonstrate the capability of SeMoP in identifying various modifications, including post-translational modifications, sequence polymorphisms, and sample handling-induced changes.
• To validate SeMoP's effectiveness in enhancing the sensitivity and accuracy of peptide modification detection in complex proteomic samples.

Main Methods:

• SeMoP employs a three-step strategy: initial protein identification via standard database search, unrestricted modification search using a novel algorithm, and targeted verification of discovered modifications.
• Utilizes low-resolution ion trap MS/MS spectra for peptide analysis.
• Incorporates decoy sequences in database searches to establish a false discovery rate (FDR).

Main Results:

• SeMoP successfully identified various sample handling-induced modifications (e.g., beta-elimination, pyrocarbamidomethylation) and biologically induced modifications (e.g., phosphorylation, methylation) in plasma proteins.
• A subsequent targeted search using SeMoP resulted in a four-fold increase in the number of identified modified peptides.
• Analysis of a cervical cancer cell line identified novel amino acid substitutions with a false discovery rate below 5% for the unrestricted search.

Conclusions:

• SeMoP is an effective and easily implemented computational approach for the discovery and verification of peptide modifications in shotgun proteomics.
• The method enhances sensitivity and accuracy, enabling the identification of a broader range of modifications.
• SeMoP holds significant potential for advancing proteomic research by providing a robust tool for comprehensive peptide modification analysis.