Quantifying the Impact of the Peptide Identification Framework on the Results of Fast Photochemical Oxidation of Protein Analysis

0583029 - MBÚ 2024 RIV US eng J - Journal Article
Zákopčaník, Marek - Kavan, Daniel - Novák, Petr - Loginov, Dmitry Sergej
Quantifying the Impact of the Peptide Identification Framework on the Results of Fast Photochemical Oxidation of Protein Analysis.
Journal of Proteome Research. Roč. 23, č. 2 (2023), s. 609-617. ISSN 1535-3893. E-ISSN 1535-3907
EU Projects: European Commission(XE) 823839 - EPIC-XS
Institutional support: RVO:61388971
Keywords : mass-spectrometry * exchange * fpop * searchengine * structural proteomics
OECD category: Microbiology
Impact factor: 4.4, year: 2022
Method of publishing: Open access
https://pubs.acs.org/doi/10.1021/acs.jproteome.3c00390

Fast Photochemical Oxidation of Proteins (FPOP) is a promising technique for studying protein structure and dynamics. The quality of insight provided by FPOP depends on the reliability of the determination of the modification site. This study investigates the performance of two search engines, Mascot and PEAKS, for the data processing of FPOP analyses. Comparison of Mascot and PEAKS of the hemoglobin--haptoglobin Bruker timsTOF data set (PXD021621) revealed greater consistency in the Mascot identification of modified peptides, with around 26% of the IDs being mutual for all three replicates, compared to approximately 22% for PEAKS. The intersection between Mascot and PEAKS results revealed a limited number (31%) of shared modified peptides. Principal Component Analysis (PCA) using the peptide-spectrum match (PSM) score, site probability, and peptide intensity was applied to evaluate the results, and the analyses revealed distinct clusters of modified peptides. Mascot showed the ability to assess confident site determination, even with lower PSM scores. However, high PSM scores from PEAKS did not guarantee a reliable determination of the modification site. Fragmentation coverage of the modification position played a crucial role in Mascot assignments, while the AScore localizations from PEAKS often become ambiguous because the software employs MS/MS merging.
Permanent Link: https://hdl.handle.net/11104/0351128