Figure 2 - Cone-like (left) and umbrella-like (right) topologies of 2-3 and 2-6 siaylated glycans.

The topology of 2-3 and 2-6 is governed by the glycosidic torsion angles of the trisaccharide motifs—Neu5Ac2-3Gal1-3/4GlcNAc and Neu5Ac2-6Gal1-4GlcNAc, respectively (Supplementary Fig. 3 online). A parameter ()—the angle between the C2 atom of Neu5Ac and C1 atoms of the subsequent Gal and GlcNAc sugars in these trisaccharide motifs—characterizes the topology. Superimposition of the contour and the conformational maps of the 2-3 and 2-6 motifs showed that the 2-3 motif adopts 100% cone-like topology and 2-6 motif sampled both the cone-like and umbrella-like topologies (Supplementary Fig. 3). In the cone-like topology sampled by 2-3 and 2-6, the GlcNAc and subsequent sugars are positioned along a region spanning a cone. The interactions of HA with the cone-like topology primarily involves contacts of amino acids at the numbered positions (based on H3 HA numbering) with Neu5Ac and Gal sugars. On the other hand, in the umbrella-like topology, which is unique to 2-6, the GlcNAc and subsequent sugars bend toward the HA binding site (as observed in HA-2-6 cocrystal structures). Longer 2-6 oligosaccharides (at least a tetrasaccharide) would favor this conformation as it is stabilized by intra-sugar van der Waals contact between acetyl groups of GlcNAc and Neu5Ac. HA interactions with sialylated glycans with the umbrella-like topology involve contacts of amino acids at the numbered positions (based on H3 HA numbering) with GlcNAc and subsequent sugars in addition to contacts with Neu5Ac and Gal sugars.