Engineering the fragment crystallizable (Fc) region of human IgG1 multimers and monomers to fine-tune interactions with sialic acid-dependent receptors.
Engineering the fragment crystallizable (Fc) region of human IgG1 multimers and monomers to fine-tune interactions with sialic acid-dependent receptors.
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DOI：
10.1074/jbc.m117.795047
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发表时间：
2017-08-04
期刊：
影响因子：
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通讯作者：
Pleass RJ
中科院分区：
文献类型：
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作者：
Blundell PA;Le NPL;Allen J;Watanabe Y;Pleass RJ
多聚体片段可结晶（Fc）区和Fc融合蛋白正被积极探索作为IVIG疗法的仿生替代物，IVIG疗法被用于管理许多疾病和病症，但昂贵且不总是有效的。人IgG 1的Fc区（IgG 1-Fc）可被工程化为多聚体结构（hexa-Fc），其以高亲合力结合其同源受体。Asn-297处连接的独特N-连接聚糖对IgG 1-Fc结构和功能的关键影响已得到充分证明;然而，N-连接聚糖是否在多聚体、亲合性结合Fc中具有类似的关键作用尚不清楚。Hexa-Fc在Asn-77（相当于IgG 1 Fc中的Asn-297）和Asn-236（相当于IgM尾段中的Asn-563）处含有两个N-连接位点。我们在这里报告，糖基化Asn-297是关键的相互作用与Fc受体和补体和糖基化Asn-563是必不可少的控制多聚化。我们还发现，在位置1的N末端引入额外的完全占据的N-连接糖基化位点（相当于IgG 1的Fc中的Asp-221）显著提高了Fc多聚体的总体唾液酸含量。此外，Cys-575在IgM尾段的多聚体的更换导致在单体与增强的唾液酸含量和差异受体结合的配置文件。因此，将额外的N-连接的聚糖插入单体或多聚体的铰链或尾段中导致具有增强的唾液酸化的分子，其可适用于控制炎症或阻断病原体入侵。
Multimeric fragment crystallizable (Fc) regions and Fc-fusion proteins are actively being explored as biomimetic replacements for IVIG therapy, which is deployed to manage many diseases and conditions but is expensive and not always efficient. The Fc region of human IgG1 (IgG1-Fc) can be engineered into multimeric structures (hexa-Fcs) that bind their cognate receptors with high avidity. The critical influence of the unique N-linked glycan attached at Asn-297 on the structure and function of IgG1-Fc is well documented; however, whether the N-linked glycan has a similarly critical role in multimeric, avidly binding Fcs, is unknown. Hexa-Fc contains two N-linked sites at Asn-77 (equivalent to Asn-297 in the Fc of IgG1) and Asn-236 (equivalent to Asn-563 in the tail piece of IgM). We report here that glycosylation at Asn-297 is critical for interactions with Fc receptors and complement and that glycosylation at Asn-563 is essential for controlling multimerization. We also found that introduction of an additional fully occupied N-linked glycosylation site at the N terminus at position 1 (equivalent to Asp-221 in the Fc of IgG1) dramatically enhances overall sialic acid content of the Fc multimers. Furthermore, replacement of Cys-575 in the IgM tail piece of multimers resulted in monomers with enhanced sialic acid content and differential receptor-binding profiles. Thus insertion of additional N-linked glycans into either the hinge or tail piece of monomers or multimers leads to molecules with enhanced sialylation that may be suitable for managing inflammation or blocking pathogen invasion.