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Quantifying the Impact of the Peptide Identification Framework on the Results of Fast Photochemical Oxidation of Protein Analysis
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SYSNO ASEP 0583029 Document Type J - Journal Article R&D Document Type Journal Article Subsidiary J Článek ve WOS Title Quantifying the Impact of the Peptide Identification Framework on the Results of Fast Photochemical Oxidation of Protein Analysis Author(s) Zákopčaník, Marek (MBU-M) ORCID
Kavan, Daniel (MBU-M) RID, ORCID
Novák, Petr (MBU-M) RID, ORCID
Loginov, Dmitry Sergej (MBU-M) RIDSource Title Journal of Proteome Research. - : American Chemical Society - ISSN 1535-3893
Roč. 23, č. 2 (2023), s. 609-617Number of pages 9 s. Language eng - English Country US - United States Keywords mass-spectrometry ; exchange ; fpop ; searchengine ; structural proteomics OECD category Microbiology Method of publishing Open access Institutional support MBU-M - RVO:61388971 UT WOS 001157569300001 EID SCOPUS 85181560169 DOI 10.1021/acs.jproteome.3c00390 Annotation 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. Workplace Institute of Microbiology Contact Eliška Spurná, eliska.spurna@biomed.cas.cz, Tel.: 241 062 231 Year of Publishing 2024 Electronic address https://pubs.acs.org/doi/10.1021/acs.jproteome.3c00390
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