Dual Glycosyltransferases from Diverge from the Canonical N-Linked Glycan Assembly Pathway.

Species within the genus are recognized as emerging human pathogens. Common to all known members of the genus is the presence of an asparagine-linked glycosylation pathway encoded by the pgl operon. species are divided into two major groups, Group I and Group II. To date, most biochemical studies have focused on the Group I species including . We recently reported that the Group II pathway deviates from that of Group I by the inclusion of a C-6″-oxidized GalNAc (GalNAcA) at the third position installed by PglJ. Herein, we investigate the diversification of the PglH enzymes that act subsequent to installation of GalNAcA. The majority of pgl operons from Group II species, including , encode two GT-B fold glycosyltransferases (GTs), PglH1 and PglH2. As the functions of these GTs were not clear by simple comparison of their sequences to that of PglH, further analyses were required. We show that subsequent to the action of PglJ, PglH2 installs the next HexNAc followed by PglH1 adding a single sugar. These steps diverge from the pathway not only in the identity of the sugar donors (UDP-GlcNAc) but also in installing single sugars rather than acting processively. These biochemical studies were extended via bioinformatics to identify sequence signatures that provide predictive capabilities for unraveling the prokaryotic glycan landscape. Phylogenetic analysis showed early divergence between the PglH orthologs and PglH1/PglH2 orthologs, leading to diversification of the final glycan.