19F NMR Monitoring of Reversible Protein Post-Translational Modifications: Class D β-Lactamase Carbamylation and Inhibition

Bacterial production of beta-lactamases with carbapenemase activity is a global health threat. The active sites of class D carbapenemases such as OXA-48, which is of major clinical importance, uniquely contain a carbamylated lysine residue which is essential for catalysis. Although there is significant interest in characterizing this post-translational modification, and it is a promising inhibition target, protein carbamylation is challenging to monitor in solution. We report the use of F-19 NMR spectroscopy to monitor the carbamylation state of F-19-labelled OXA-48. This method was used to investigate the interactions of OXA-48 with clinically used serine beta-lactamase inhibitors, including avibactam and vaborbactam. Crystallographic studies on F-19-labelled OXA-48 provide a structural rationale for the sensitivity of the F-19 label to active site interactions. The overall results demonstrate the use of F-19 NMR to monitor reversible covalent post-translational modifications.