Characterisation of the Dynamic Interactions between Complex N‐Glycans and Human CD22

Abstract

CD22 (Siglec‐2) is a B‐cell surface inhibitory protein capable of selectively recognising sialylated glycans, thus dampening autoimmune responses against self‐antigens. Here we have characterised the dynamic recognition of complex‐type N‐glycans by human CD22 by means of orthogonal approaches including NMR spectroscopy, computational methods and biophysical assays. We provide new molecular insights into the binding mode of sialoglycans in complex with h‐CD22, highlighting the role of the sialic acid galactose moieties in the recognition process, elucidating the conformational behaviour of complex‐type N‐glycans bound to Siglec‐2 and dissecting the formation of CD22 homo‐oligomers on the B‐cell surface. Our results could enable the development of additional therapeutics capable of modulating the activity of h‐CD22 in autoimmune diseases and malignancies derived from B‐cells.

Unveiling Molecular Recognition of Sialoglycans by Human Siglec-10

“…Here, the Neu5Ac-a-(2-6)-Gal glycosidic linkage was defined by an extra torsion angle (Poppe et al, 1992), namely, u (O 0 6 ÀC 0 6 ÀC 5 'ÀO 0 5 ), that influences the entire three-dimensional structure of 2. In detail, three different rotamers differing in the u value can coexist in solution, namely, gg/tg/gt (Patel et al, 2014) (u À60 /180 /60 , respectively), the gt conformer being the most populated in the free state (Di Carluccio et al, 2019). Comparing the experimental NOE-derived and theoretical distances (Table 2, Figure S2 right panel), a preference was evident for the geometry with 4/c/u torsion angles of À60 /180 /60 around Sia-Gal linkage (Family I, gt conformer) (Poppe et al, 1992) both in the free and bound states.…”

Molecular Recognition Insights of Sialic Acid Glycans by Distinct Receptors Unveiled by NMR and Molecular Modeling

“…The representative structures were generated using the Carbohydrate Builder tool from GLYCAM-web (Woods, 2005), and the images were created using PyMOL 2.4.1 (Schrödinger). the H5 of Neu5Ac and the NHAc of GlcNAc indicates that the conformer holding φ around −60 °is the major conformer in solution (Sassaki et al, 2013;Forgione et al, 2020a;Di Carluccio et al, 2020). Additionally, the value of the ω dihedral angle that can adopt −60 °/180 °/60 °corresponding to gg/tg/gt rotamers, respectively, has shown values around 60 °(gt conformer) (Di Carluccio et al, 2019) (Table 2).…”

“…The ψ dihedral angle (C2-O-C3′-H3′) remains stable at around 180 °. The information about each conformer and the corresponding NMR evidence proving their existence was retrieved from Forgione et al (2020a) and Di Carluccio et al (2020). The representative structures were generated using the Carbohydrate Builder tool from GLYCAM-web (Woods, 2005), and the images were created using PyMOL 2.4.1 (Schrödinger).…”

Molecular recognition of sialoglycans by streptococcal Siglec-like adhesins: toward the shape of specific inhibitors

“…The N-terminal V-set Ig domain in the Siglec family contains common structural determinants, such as a conserved arginine in the F strand, key residue for the formation of salt bridge with the carboxyl group of sialic acid. 22,35,36 As for the Siglec-like adhesins SLBR-H and SLBR-B, despite the proximity in the binding site of arginine residues (Arg340 SLBR-H and Arg484 SLBR-B ) to the sialic acid unit of the ligands, no similar ionic interactions with the carboxyl group of Neu5Ac were detected, but important hydrogen bonds were found with the glycerol moiety, in particular with O8 and O9 of Neu5Ac. Instead, the carboxylate group of Neu5Ac established interactions with the threonine residue of the ΦTRY motif (Thr339 SLBR-H and Thr483 SLBR-B ).…”