A chemoenzymatic method for simultaneous profiling N- and O-glycans on glycoproteins using one-pot format.
Cell Rep Methods
; : 100834, 2024 Aug 05.
Article
em En
| MEDLINE
| ID: mdl-39116882
ABSTRACT
Glycosylation is generally characterized and controlled as a critical quality attribute for therapeutic glycoproteins because glycans can impact protein drug-product efficacy, half-life, stability, and safety. Analytical procedures to characterize N-glycans are relatively well established, but the characterization of O-glycans is challenging due to the complex workflows and lack of enzymatic tools. Here, we present a simplified chemoenzymatic method to simultaneously profile N- and O-glycans from the same sample using a one-pot format by mass spectrometry (MS). N-glycans were first released by PNGase F, followed by O-glycopeptide generation by proteinase K, selective N-glycan reduction, and O-glycan release by ß-elimination during permethylation of both N- and O-glycans. Glycan structural assignments and determination of N- to O-glycan ratio was obtained from the one-pot mass spectra. The streamlined, one-pot method is a reliable approach that will facilitate advanced characterizations for quality assessments of therapeutic glycoproteins.
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1
Coleções:
01-internacional
Base de dados:
MEDLINE
Idioma:
En
Revista:
Cell Rep Methods
Ano de publicação:
2024
Tipo de documento:
Article