Human noroviruses (HuNoV), members of the family Caliciviridae, are the major cause of acute viral gastroenteritis worldwide. Successful infection is linked to the ability of the protruding (P) domain of the viral capsid to bind histo-blood group antigens (HBGA). Binding to gangliosides plays a major role for many nonhuman calici- and noroviruses. Increasing evidence points to a broader role of sialylated carbohydrates such as gangliosides in norovirus infection. Here, we compare HBGA and ganglioside binding of a GII.4 HuNoV variant (MI001), previously shown to be infectious in a HuNoV mouse model. Saturation transfer difference nuclear magnetic resonance spectroscopy, native mass spectrometry (MS) and surface plasmon resonance spectroscopy were used to characterize binding epitopes, affinities, stoichiometry and dynamics, focusing on 3′-sialyllactose, the GM3 ganglioside saccharide and B antigen. Binding was observed for 3′-sialyllactose and various HBGAs following a multistep binding process. Intrinsic affinities (Kd) of fucose, 3′-sialyllactose and B antigen were determined for the individual binding steps. Stronger affinities were observed for B antigen over 3′-sialyllactose and fucose, which bound in the mM range. Binding stoichiometry was analyzed by native MS showing the presence of four B antigens or two 3′-sialyllactose in the complex. Epitope mapping of 3′-sialyllactose revealed direct interaction of α2,3-linked sialic acid with the P domain. The ability of HuNoV to engage multiple carbohydrates emphasizes the multivalent nature of norovirus glycan-specificity. Our findings reveal direct binding of a GII.4 HuNoV P dimer to α2,3-linked sialic acid and support a broader role of ganglioside binding in norovirus infection.

中文翻译：

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人诺如病毒GII.4（MI001）P二聚体结合岩藻糖基化和唾液酸化碳水化合物

人诺如病毒（HuNoV），杯状病毒科成员，是全世界急性病毒性胃肠炎的主要原因。成功感染与病毒衣壳突出（P）域结合组织血型组抗原（HBGA）的能力有关。对神经节苷脂的结合在许多非人类杯状和诺如病毒中起着重要作用。越来越多的证据表明，唾液酸化的碳水化合物（例如神经节苷脂）在诺如病毒感染中具有更广泛的作用。在这里，我们比较了先前显示在HuNoV小鼠模型中具有传染性的GII.4 HuNoV变体（MI001）的HBGA和神经节苷脂结合。饱和转移差异核磁共振波谱，天然质谱（MS）和表面等离子体共振谱用于表征结合表位，亲和力，化学计量和动力学，重点研究3'-唾液乳糖，GM3神经节苷脂和B抗原。在多步结合过程之后，观察到3'-唾液乳糖和各种HBGA的结合。对于各个结合步骤，确定了岩藻糖，3'-唾液乳糖和B抗原的内在亲和力（Kd）。观察到B抗原比3'-唾液乳糖和岩藻糖具有更强的亲和力，后者在mM范围内结合。通过天然MS分析结合化学计量，显示复合物中存在四个B抗原或两个3'-唾液乳糖。3'-唾液乳糖的表位作图揭示了α2,3-连接的唾液酸与P结构域的直接相互作用。HuNoV参与多种碳水化合物的能力强调了诺如病毒聚糖特异性的多价性质。我们的发现揭示了GII.4 HuNoV P二聚体与α2,3-连接的唾液酸的直接结合，并支持神经节苷脂结合在诺如病毒感染中发挥更广泛的作用。