Unlike Natural Killer (NK) p30, Natural Cytotoxicity Receptor NKp44 Binds to Multimeric α2,3-NeuNAc-Containing N-Glycans
Unlike Natural Killer (NK) p30, Natural Cytotoxicity Receptor NKp44 Binds to Multimeric α2,3-NeuNAc-Containing N-Glycans
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DOI：
10.1248/bpb.35.594
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发表时间：
2012-04-01
影响因子：
2
通讯作者：
Matsumoto, Kojiro
中科院分区：
文献类型：
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作者：
Ito, Kenichiro;Higai, Koji;Matsumoto, Kojiro
天然细胞毒受体2(NCR2或自然杀伤细胞(NK)p44)和NCR3(NKp30)与肝素和硫酸肝素结合；然而，其他天然配体尚未确定。我们以前报道过NCR1(NKp46)可以与含有NeuNAc的多聚体N-糖链和硫酸糖链结合。在这项研究中，我们利用NKp44-H6和NKp30-H6标记的共同重组胞外区(NKp44-H6和NKp30-H6)，研究了NKp44和NKp30是否能与含有NeuNAc的糖链结合。NKp44-H6，而不是NKp30-H6，结合了表达转铁蛋白的多聚体唾液酸化Lewis X，其K-d为420 nm。竞争结合和直接结合分析表明，NKp44-H6主要识别N-糖链非还原末端的α-2，3-NeuNAc残基。此外，NKp44-H6的定点突变体，如R47Q、R55Q、R92Q、R95Q、K103Q和R106Q，都减少了与α-2，3-唾液酸化N-糖链的结合。这些结果表明，NKp44通过离子相互作用与α-2，3-唾液酸化的N-糖链结合，并且这些结合部位可能与肝素结合部位有一定的重叠。
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 6xHis (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 alpha 2,3-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 alpha 2,3-sialylated N-glycans. These results suggest that NKp44 binds to alpha 2,3-sialylated N-glycans through ionic interactions, and that these binding sites might have some overlap with heparin binding sites.