Human noroviruses (HuNoVs) are a major cause of acute gastroenteritis. Many HuNoVs recognize histo-blood group antigens (HBGAs) as cellular receptors or attachment factors for infection. It was recently proposed that HuNoV recognition of HBGAs involves a cooperative, multistep binding mechanism that exploits both known and previously unknown glycan binding sites. In this study, binding measurements, implemented using electrospray ionization mass spectrometry (ESI-MS) were performed on homodimers of the protruding domain (P dimers) of the capsid protein of three HuNoV strains [Saga (GII.4), Vietnam 026 (GII.10) and VA387 (GII.4)] with the ethyl glycoside of the B trisaccharide (α-d-Gal-(1→3)-[α-l-Fuc-(1→2)]-β-d-Gal-OC₂H₅) and free B type 1 tetrasaccharide (α-d-Gal-(1→3)-[α-l-Fuc-(1→2)]-β-d-Gal-(1→3)-d-GlcNAc) in an effort to confirm the existence of new HBGA binding sites. After correcting the mass spectra for nonspecific interactions that form in ESI droplets as they evaporate to dryness, all three P dimers were found to bind a maximum of two B trisaccharides at the highest concentrations investigated. The apparent affinities measured for stepwise binding of B trisaccharide suggest positive cooperativity. Similar results were obtained for B type 1 tetrasaccharide binding to Saga P dimer. Based on these results, it is proposed that HuNoV P dimers possess only two HBGA binding sites. It is also shown that nonspecific binding corrections applied to mass spectra acquired using energetic ion source conditions that promote in-source dissociation can lead to apparent HuNoV–HBGA oligosaccharide binding stoichiometries and affinities that are artificially high. Finally, evidence that high concentrations of oligosaccharide can induce conformational changes in HuNoV P dimers is presented.

中文翻译：

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定量组织血型组抗原寡糖对人诺如病毒的结合化学计量和亲和力

人类诺如病毒（HuNoVs）是急性胃肠炎的主要原因。许多HuNoV都将组织血型抗原（HBGA）视为感染的细胞受体或附着因子。最近有人提出，对HBGA的HuNoV识别涉及一种合作的多步结合机制，该机制利用了已知的和以前未知的聚糖结合位点。在这项研究中，对三种HuNoV株[Saga（GII.4），越南026（GII） 0.10）和VA387（GII.4）]用B三糖的乙基糖苷（α- d -Gal-（1→3） - [α-升-Fuc-（1→2）] - β- d - Gal-OC ₂ H ₅）和游离的B型1四糖（α- d -Gal-（1→3）-[α- 1- Fuc-（1→2）]-β- d- Gal-（1→3）-d-GlcNAc），以确认是否存在新的HBGA结合位点。在校正了ESI液滴蒸发至干时形成的非特异性相互作用的质谱图后，发现所有三个P二聚体在所研究的最高浓度下最多可结合两个B三糖。逐步亲和力B三糖的表观亲和力表明积极合作。对于B型1四糖与Saga P二聚体的结合，获得了相似的结果。基于这些结果，提出HuNoV P二聚体仅具有两个HBGA结合位点。研究还表明，使用能促进源内解离的高能离子源条件对质谱进行的非特异性结合校正可以导致明显的HuNoV-HBGA寡糖结合化学计量比和亲和力过高。最后，提出了高浓度低聚糖可诱导HuNoV P二聚体构象变化的证据。