Ubiquitin is a carbon dioxide-binding protein

The identification of CO₂-binding proteins is crucial to understanding CO₂-regulated molecular processes. CO₂ can form a reversible posttranslational modification through carbamylation of neutral N-terminal α-amino or lysine ε-amino groups. We have previously developed triethyloxonium (TEO) ion as a chemical proteomics tool for covalent trapping of carbamates, and here, we deploy TEO to identify ubiquitin as a mammalian CO₂-binding protein. We use ¹³C-NMR spectroscopy to demonstrate that CO₂ forms carbamates on the ubiquitin N terminus and ε-amino groups of lysines 6, 33, 48, and 63. We demonstrate that biologically relevant pCO₂ levels reduce ubiquitin conjugation at lysine-48 and down-regulate ubiquitin-dependent NF-κB pathway activation. Our results show that ubiquitin is a CO₂-binding protein and demonstrates carbamylation as a viable mechanism by which mammalian cells can respond to fluctuating pCO₂.