Quantitative O-GlcNAc analysis between wild type and sec-5 using SILIA (F1 on Eclipse (EThcD)

O-GlcNAcylation is a crucial post-translational modification of proteins observed in both plants and animals and plays a key role in growth and development. Plants have close OGT homologs, SECRET AGENT (SEC) and SPINDLY (SPY). In vitro, SEC has shown O-GlcNAc activity. Recently, a surprising discovery in wheat revealed an atypical TaOGT(TaOGT1) with no structural similarity to SEC and SPY enzyme (Fan et al. 2021). TaOGT1 was found to O-GlcNAcylate TaGRP2. In Arabidopsis, approximately 34 unannotated or uncharacterized proteins share similarity with TaOGT1, leading to questions about whether SEC is the primary contributor to O-GlcNAcylation in plants. Here we use LWAC enrichment and SILIA labeling, quantifying at both MS1. Our findings reveal a significant reduction in O-GlcNAc levels in the sec mutant, indicating SEC’s critical role in mediating O-GlcNAcylation.