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Abstract
The bacterial cell wall serves as a protective barrier that encapsulates the cell and features an array of macromolecules, including polysaccharides. These sugar polymers form an extracellular coating that is crucial for bacterial survival and can contribute to infections in host organisms. Bacterial glycans are biosynthesized in a process that is typically initiated by an important class of enzymes called phosphoglycosyltransferases (PGTs). PGTs are integral membrane proteins that are responsible for the first committed step in bacterial glycan biosynthesis, which involves catalyzing the transfer of a phosphosugar moiety to a membrane-embedded polyprenyl phosphate substrate. Despite playing an important role in this process, a lack of information remains regarding PGT function and the reactions they catalyze. One approach to studying the function of enzymes involves probing their substrate specificities using small molecules with varying chemical functionalities. Herein, a small library of short-chain isoprenoid analogues with bulky aromatic groups was developed to probe the effects of substrate size on PGT function. Seven neryl phosphate analogues with naphthylamine, aminobiphenyl, and benzylaniline incorporated were utilized in chemoenzymatic reactions to study the monotopic PGTs, PglC (Campylobacter jejuni) and WcfS (Bacteroides fragilis). It was found that all seven synthetic probes were accepted by PglC, while only two were accepted by WcfS. This was an important development as short-chain analogues previously tested against WcfS were ineffective suggesting that small structural changes in the substrate could impact activity.