Simple and practical sialoglycan encoding system reveals vast diversity in nature and identifies a universal sialoglycan-recognizing probe derived from AB5 toxin B subunits

Vertebrate sialic acids (Sias) display much diversity in modifications, linkages and underlying glycans. Slide microarrays allow high-throughput explorations of sialoglycan-protein interactions. A microarray presenting ~150 structurally-defined sialyltrisaccharides with various Sias linkages and modifications still poses challenges in planning, data sorting, visualization and analysis. To address these issues, we devised a simple 9- digit code for sialyltrisaccharides with terminal Sias and underlying two monosaccharides assigned from the non- reducing end, with three digits assigning a monosaccharide, its modifications, and linkage. Calculations based on the encoding system reveals >113,000 likely linear sialyltrisaccharides in nature. Notably a biantennary N-glycan with two terminal sialyltrisaccharides could thus have >1010 potential combinations and a triantennary N-glycan with three terminal sequences, >1015 potential combinations. While all possibilities likely do not exist in nature, sialoglycans encode enormous diversity. While glycomic approaches are used to probe such diverse sialomes, naturally-occurring bacterial AB5 toxin B subunits are simpler tools to track the dynamic sialome in biological systems. Sialoglycan microarray was utilized to compare sialoglycan-recognizing bacterial toxin B subunits. Unlike the poor correlation between B subunits and species phylogeny, there is stronger correlation with Sia- epitope preferences. Further supporting this pattern, we report a B subunit (YenB) from Yersinia enterocolitica (broad host range) recognizing almost all sialoglycans in the microarray, including 4-O-acetylated-Sias not recognized by a Y. pestis orthologue (YpeB). Differential Sia-binding patterns were also observed with phylogenetically-related B subunits from Escherichia coli (SubB), Salmonella Typhi (PltB), S. Typhimurium (ArtB), extra-intestinal E.coli (EcPltB), Vibrio cholera (CtxB), and cholera family homologue of E. coli (EcxB).