Expanding the Chemoproteomic Toolkit to Asparagine and Glutamine

Chemoproteomic strategies have revolutionized proteome
annotation
by targeting nucleophilic and redox-active side chains. However, the
primary amides of asparagine (Asn) and glutamine (Gln) have long lacked
robust chemical tools for proteome-wide interrogation. We report a
chemoselective palladium-mediated dehydration that converts Asn/Gln
amides to nitriles under mild aqueous conditions. This transformation
enables the first proteome-wide mapping of chemically addressable
Asn/Gln sites in lysates and living cells. Leveraging this reactivity,
we establish an inverse chemoproteomic framework in which reduced
nitrile formation reports PTM-mediated protection of Asn/Gln sites,
including those impacted by deamidation and N-glycosylation. This
approach reveals sites masked by post-translational modifications
(PTMs), specifically those associated with deamidation and N-glycosylation.
In yeast, this framework expanded the known N-glycoproteome, identifying
numerous candidates missed by traditional glycopeptide enrichment
due to low abundance or noncanonical motifs. Furthermore, comparative
profiling in Candida albicans captured
the dynamic remodeling of glycosylation patterns during morphogenesis.
This dehydration-to-nitrile platform establishes a scalable handle
on the amide proteome to map residue accessibility and PTM-linked
site dynamics across biological states.