- Titel
- A Workflow towards the Reproducible Identification and Quantitation of Protein Carbonylation Sites in Human Plasma
- Zitierfähige Url:
- https://nbn-resolving.org/urn:nbn:de:bsz:15-qucosa2-849111
- Quellenangabe
- Antioxidants
Erscheinungsjahr: 2021
Jahrgang: 10
Heft: 3
E-ISSN: 2076-3921
Artikelnummer: 369 - Erstveröffentlichung
- 2021
- Abstract (EN)
- Protein carbonylation, a marker of excessive oxidative stress, has been studied in the context of multiple human diseases related to oxidative stress. The variety of post-translational carbonyl modifications (carbonyl PTMs) and their low concentrations in plasma challenge their reproducible identification and quantitation. However, carbonyl-specific biotinylated derivatization tags (e.g., aldehyde reactive probe, ARP) allow for targeting carbonyl PTMs by enriching proteins and peptides carrying these modifications. In this study, an oxidized human serum albumin protein model (OxHSA) and plasma from a healthy donor were derivatized with ARP, digested with trypsin, and enriched using biotin-avidin affinity chromatography prior to nano reversed-phase chromatography coupled online to electrospray ionization tandem mass spectrometry with travelling wave ion mobility spectrometry (nRPC-ESI-MS/MS-TWIMS). The presented workflow addresses several analytical challenges by using ARP-specific fragment ions to reliably identify ARP peptides. Furthermore, the reproducible recovery and relative quantitation of ARP peptides were validated. Human serum albumin (HSA) in plasma was heavily modified by a variety of direct amino acid oxidation products and adducts from reactive carbonyl species (RCS), with most RCS modifications being detected in six hotspots, i.e., Lys10, Lys190, Lys199, Lys281, Lys432, and Lys525 of mature HSA.
- Andere Ausgabe
- Link zur Erstveröffentlichung
Link: https://doi.org/10.3390/antiox10030369 - Freie Schlagwörter (EN)
- protein carbonylation; human plasma; aldehyde reactive probe (ARP); biotin-avidin affinity; LC-MS/MS
- Klassifikation (DDC)
- 540
- Verlag
- MDPI, Basel
- Version / Begutachtungsstatus
- publizierte Version / Verlagsversion
- URN Qucosa
- urn:nbn:de:bsz:15-qucosa2-849111
- Veröffentlichungsdatum Qucosa
- 24.04.2023
- Dokumenttyp
- Artikel
- Sprache des Dokumentes
- Englisch
- Lizenz / Rechtehinweis
CC BY 4.0