Affinity-Based Covalent Sialyltransferase Probes Enabled by Ligand-Directed Chemistry

Sialyltransferases (ST) are enzymes found in among others mammals and bacteria that are responsible for producing sialylated glycans, which are known to play important roles in human health and disease. An example of the latter being the opportunistic molecular mimicry by pathogenic bacteria. However, chemical tools to study sialyltransferases have been limited to non-covalent inhibitors and probes that do not allow isolation and profiling of these important enzymes. Here we report the first ever covalent probes for ST by utilising ligand-directed chemistry. These affinity-based probes are armed with a simple to synthesise but robust O-nitrobenzoxadiazole (O-NBD) warhead, which is a lysine specific SNAr electrophilic warhead with a desirable turn-on fluorescence property. We demonstrate their high specificity when applied to label both recombinant sialyltransferases as well as native lipooligosaccharide sialyltransferase (Lst) in Neisseria gonorrhoeae, a relevant human pathogen. This new class of modular covalent ST probes, and their future iterations, could pave the way for new detailed studies of sialyltransferases in their native environment.