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Cited by 0Advanced Proteomics Techniques and ApplicationsUbiquitin and proteasome pathwaysMass Spectrometry Techniques and Applications

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Improved Peptide Search for Identification of SUMO and Sequence-Based Modifiers, in MaxSBM

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Lennartsson, Caroline, Kyriakidou, Pelagia, Nielsen, Michael Lund, Olsen, Jesper Velgaard, Cox, Jürgen, Hendriks, Ivo Alexander·2026·Molecular & Cellular Proteomics
DOI 10.1016/j.mcpro.2026.101589PMC PMC13292685

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Abstract

Abstract Post-translational modifications (PTMs), such as SUMOylation and ubiquitination, regulate key cellular processes by covalently attaching to lysine residues. While mass spectrometry allows site-specific identification of PTMs, most existing search engines are optimized for small, non-fragmenting modifications and struggle to detect large, fragmenting protein-based modifiers. We refer to these as Sequence-Based Modifiers (SBMs). To overcome this limitation, we developed an SBM-specific search strategy within MaxQuant that accounts for the fragmentation behavior of SBMs during peptide identification. Using publicly available datasets, we validated our approach for SUMO2/3. Our analysis identified distinct diagnostic features and characteristic mass shifts associated with SBM fragmentation, referred to in this study as d-ions (diagnostic ions) and p-ions. By leveraging these features, our method improved the identification of SUMOylated peptides from human cell lines by 13%, SUMOylation sites in mouse embryonic cells by 18%, and in mouse adipocytes by 25%. Our search method improved spectral annotation of SBMs by up to 9% increase in the median Andromeda score. Taken together, we highlight the potential of our SBM search to enhance the discovery of protein-based modifications. Highlights Development of a MaxQuant module tailored for identifying Sequence-Based Modifiers (SBMs), including SUMO2/3 Incorporation of SBM-specific fragmentation patterns into search algorithms Enhanced biological discovery through improved PTM identification from mass spectrometry datasets In Brief Here, we introduce MaxSBM, an optimized framework for interpreting complex sequence-based modifiers (SBMs), particularly SUMO, within MaxQuant. Our approach incorporates SBM-specific d- and p-ion series into peptide scoring and annotation. By extending the theoretical spectral space to include fragments bearing partial SUMO (or other SBM) peptide remnants, MaxSBM provides a more comprehensive spectral annotation which enhances peptide scoring, resulting in increased identification rates at a higher confidence. Beyond methodological refinement, we validated MaxSBM via reanalysis of several physiological SUMO datasets, ultimately unlocking new insights via mapping of previously obscured modification sites.

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Metadata

Authors
Lennartsson, C., Kyriakidou, P., Nielsen, M. L., Olsen, J. V., Cox, J., & Hendriks, I. A.
Affiliations
  • University of Copenhagen (DK)
  • Max Planck Institute of Biochemistry (DE)
Corresponding author
Hendriks, Ivo Alexander
ORCID
Venue
Molecular & Cellular Proteomics
Publisher
Elsevier BV
Published
2026-06
Volume
25
Issue
6
Pages
101589
ISSN
1535-9476 (ISSN-L)
DOI
10.1016/j.mcpro.2026.101589
PMC
PMC13292685
OpenAlex
W4413900229
Cited by
0
References
2
FWCI
0.00
Type
journal article
Language
en
Open access
Licence
Locations
  • Molecular & Cellular Proteomics (publishedVersion, cc-by-nc-nd)
  • bioRxiv (Cold Spring Harbor Laboratory) (acceptedVersion, cc-by-nc-nd)
  • PubMed Central (publishedVersion, cc-by-nc-nd)
  • PubMed (publishedVersion)
  • Research at the University of Copenhagen (University of Copenhagen) (publishedVersion, cc-by-nc-nd)
  • MPG.PuRe (Max Planck Society) (submittedVersion)
Funders
  • Novo Nordisk Foundation Center for Protein Research (NNF24SA0098829)
  • Danish Cancer Society (R146-A9159-16-S2)
  • Danmarks Frie Forskningsfond (8020-00220B, 0135-00096B)
  • Novo Nordisk Fonden
  • Horizon 2020 Framework Programme (EPIC-XS-823839)
Topics
  • Physical Sciences › Chemistry › Spectroscopy › Advanced Proteomics Techniques and Applications
  • Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › Ubiquitin and proteasome pathways
  • Physical Sciences › Chemistry › Spectroscopy › Mass Spectrometry Techniques and Applications
Keywords
SUMOylation, MaxQuant, peptide identification, post-translational modification, SUMO2/3, fragmentation patterns, diagnostic ions, spectral annotation
MeSH terms
Amino Acid Sequence, Animals, Humans, Peptides, Protein Processing, Post-Translational, Ubiquitins, Adipocytes, Small Ubiquitin-Related Modifier Proteins, Proteomics, Mice, Sumoylation

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