ArticlePloS one2021
Evaluation of mass spectrometry MS/MS spectra for the presence of isopeptide crosslinked peptides.
Article in PloS one, 2021. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Cited by 7 papers.
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Who cites it
7 citing papers in PubMed, 9 citations in OpenAlex.
- Enzymatic Encoding of Topology in an Intrinsically Disordered Single-Chain Protein.Angewandte Chemie (International ed. in English) · 2026Article
- Asymmetric isopeptide bond steers directional genomic RNA egress from icosahedral virus.Science advances · 2025Article
- Perforated imprinting on high moisture meat analogue confers long range mechanical anisotropy resembling meat cuts.NPJ science of food · 2024Article
- Dichlorvos-induced formation of isopeptide crosslinks between proteins in SH-SY5Y cells.Analytical biochemistry · 2022Article
- Organophosphorus Pesticides Promote Protein Cross-Linking.Chemical research in toxicology · 2022Article
- Mass Spectrometric Identification of a Novel Factor XIIIa Cross-Linking Site in Fibrinogen.Proteomes · 2021Article
- Rabbit Antidiethoxyphosphotyrosine Antibody, Made by Single B Cell Cloning, Detects Chlorpyrifos Oxon-Modified Proteins in Cultured Cells and Immunopurifies Modified Peptides for Mass Spectrometry.Journal of proteome research · 2021Article
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Authors and funding
3 authors at 1 institution in 2 countries.
Funding
Abstract
Isopeptide crosslinked proteins can be the product of transglutaminase or of exposure to organophosphorus toxicants (OP). Transglutaminase links glutamine to lysine with loss of ammonia. OP toxicants induce a link between glutamic acid and lysine with loss of water. Our goal was to establish criteria to distinguish real from false isopeptide crosslinks reported by software searches of mass spectrometry data. We used fragmentation spectra of tryptic peptides from MAP-rich tubulin Sus scrofa as a test system for detection of naturally-occurring isopeptide crosslinks. Data were analyzed with Protein Prospector. Criteria for the assignments included the presence of at least 1 crosslink specific product ion, fragment ions from both peptides, Protein Prospector scores ≥20, and best fit of the MS/MS data to the crosslinked peptide as opposed to a linear peptide. Out of 301,364 spectra, 15 potential transglutaminase-type crosslinked peptide candidates were identified. Manual evaluation of these MS/MS spectra reduced the number to 1 valid crosslink between Q112 of NFH and K368 of Tau. Immunopurification with anti-isopeptide 81D1C2 confirmed that MAP-rich tubulin contained only one isopeptide. Support for this isopeptide bond was obtained by showing that transglutaminase was capable of incorporating dansyl-aminohexyl -QQIV into K368. A model of the KIETHK-QLEAHNR isopeptide was synthesized with the aid of transglutaminase. MS/MS spectra of the model validated our interpretation of the native isopeptide. An OP-induced isopeptide bond between K163 of tubulin alpha-1A and E158 of tubulin beta-4B was induced by treating MAP-rich tubulin with 100 μM chlorpyrifos oxon. This crosslink was supported by the criteria described above and by the presence of diethoxyphospho-lysine 163 in the tubulin alpha-1A peptide. The information obtained in this work is valuable for future studies that aim to understand why exposure to OP is associated with increased risk of neurodegenerative disease.
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