title: Unlike natural killer (NK) p30, natural cytotoxicity receptor NKp44 binds to multimeric α2,3-NeuNAc-containing N-glycans.
abstract: Natural cytotoxicity receptor 2 (NCR2 or natural killer (NK)p44) and NCR3 (NKp30) bind to heparin and heparin sulfate; however, other natural ligands have yet to be identified. We previously reported that NCR1 (NKp46) can bind to multimeric NeuNAc-containing N-glycans and sulfated glycans. In this study, we investigated whether NKp44 and NKp30 can bind to NeuNAc-containing glycans using their common recombinant extracellular domain tagged with 6×His (NKp44-H6 and NKp30-H6). NKp44-H6, but not NKp30-H6, bound multimeric sialyl Lewis X expressing transferrin secreted by HepG2 cells (HepTF) with a K(d) of 420 nM. Competitive and direct binding assays revealed that NKp44-H6 mainly recognizes α2,3-NeuNAc residues on non-reducing ends of N-glycans on HepTF. Moreover, site-directed mutants of NKp44-H6, such as R47Q, R55Q, R92Q, R95Q, K103Q, and R106Q, had reduced binding to α2,3-sialylated N-glycans. These results suggest that NKp44 binds to α2,3-sialylated N-glycans through ionic interactions, and that these binding sites might have some overlap with heparin binding sites.
authors.affiliation: Department of Clinical Chemistry, Faculty of Pharmaceutical Sciences, Toho University, Funabashi, Chiba, Japan.
authors.forename: Kenichiro
authors.initials: K
authors.lastname: Ito
authors.forename: Koji
authors.initials: K
authors.lastname: Higai
authors.forename: Chihiro
authors.initials: C
authors.lastname: Shinoda
authors.forename: Mizuha
authors.initials: M
authors.lastname: Sakurai
authors.forename: Kazuyuki
authors.initials: K
authors.lastname: Yanai
authors.forename: Yutaro
authors.initials: Y
authors.lastname: Azuma
authors.forename: Kojiro
authors.initials: K
authors.lastname: Matsumoto
mesh_terms: D056945:Hep G2 Cells; D006493:Heparin / Q000378:metabolism; D006801:Humans; D055629:Natural Cytotoxicity Triggering Receptor 2 / Q000378:metabolism*; D055628:Natural Cytotoxicity Triggering Receptor 3 / Q000378:metabolism; D009844:Oligosaccharides / Q000378:metabolism; D011134:Polysaccharides / Q000378:metabolism*; D012710:Serum Albumin, Bovine / Q000378:metabolism; D000080506:Sialyl Lewis X Antigen
chemical_list: C112837:NCR2 protein, human; C431710:NCR3 protein, human; D055629:Natural Cytotoxicity Triggering Receptor 2; D055628:Natural Cytotoxicity Triggering Receptor 3; D009844:Oligosaccharides; D011134:Polysaccharides; D000080506:Sialyl Lewis X Antigen; D012710:Serum Albumin, Bovine; D006493:Heparin