Siglecs, sialic-acid-binding immunoglobulin-like lectins, are key immune cell receptors that recognize sialic acid residues on cell surfaces. Pathogens and tumor cells exploit Siglecs to evade immune responses and modulate immunity, contributing significantly to infectious disease and cancer pathogenesis. Siglec-7, primarily expressed on natural killer (NK) cells, functions as an inhibitory receptor, tightly regulating the immune activity. This study investigates the interaction between Siglec-7 and the capsular polysaccharide (CPS) of Neisseria meningitidis serogroup Y (Men-Y), a bacterium whose sialylated CPS is critical for virulence. We demonstrate that Men-Y CPS binds to inhibitory Siglec-7, potentially dampening immune recognition. We employed a multifaceted approach, combining biochemical and biophysical techniques to dissect this interaction. Enzyme-linked immunosorbent assays (ELISAs) and fluorescence titrations quantified the binding specificity and affinity. Ligand- and protein-based nuclear magnetic resonance (NMR) spectroscopy, coupled with computational modeling, provides detailed molecular insights. We highlight the critical influence of the Men-Y CPS conformation and sialic acid presentation on Siglec-7 binding. The specific arrangement of α-2,6-linked sialic acids on the CPS is crucial for Siglec-7 binding, demonstrating the importance of the CPS 3D structure. Preliminary immunological assays using stimulated U937 cells (a promonocytic cell line) further support the immunomodulatory role of Siglec-7 mediated by Men-Y CPS. These results offer valuable insights into the development of targeted therapeutic strategies against bacterial infections.

Department of Biochemistry Faculty of Science Charles University Hlavova 2030 8 12800 Prague Czech Republic

Department of Chemical Sciences University of Naples Federico 2 via Cinthia 4 80126 Naples Italy

Department of Molecular Cell Biology and Immunology Amsterdam UMC Vrije Universiteit Amsterdam Amsterdam 1081 HV The Netherlands

Department of Molecular Medicine and Medical Biotechnology University of Naples Federico 2 via Pansini 5 80131 Naples Italy

Finlay Vaccine Institute 21st Ave N◦ 19810 between 198 and 200 St Atabey Playa 10400 Havana Cuba

Institute of Biomolecular Chemistry National Research Council via Campi Flegrei 34 80078 Pozzuoli Naples Italy

Magnetic Resonance Centre CIRMMP and Department of Chemistry Ugo Schiff University of Florence 50019 Sesto Fiorentino Italy

Stem Cell Biology Laboratory Department of Biology University of Naples Federico 2 80131 Naples Italy

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