VHH stands for the variable regions of heavy chain only of camelid IgGs. The VHH family forms a set of interesting proteins derived from antibodies that maintain their capacity to recognize the antigen, despite their relatively small molecular weight (in the 12,000 Da range). Continuing our exploration of the possibilities of those molecules, we chose to design alternative molecules with maintained antigen recognition, but enhanced capacity, by fusing four VHH with one Fc, the fragment crystallizable region of antibodies. In doing so, we aimed at having a molecule with superior quantitative antigen recognition (×4) while maintaining its size below the 110 kDa. In the present paper, we described the building of those molecules that we coined VHH₂-Fc-VHH₂. The structure of VHH₂-Fc-VHH₂ in complex with HER2 antigen was determined using electronic microscopy and modeling. The molecule is shown to bind four HER2 proteins at the end of its flexible arms. VHH₂-Fc-VHH₂ also shows an internalization capacity via HER2 receptor superior to the reference anti-HER2 monoclonal antibody, Herceptin®, and to a simple fusion of two VHH with one Fc (VHH₂-Fc). This new type of molecules, VHH₂-Fc-VHH₂, could be an interesting addition to the therapeutic arsenal with multiple applications, from diagnostic to therapy.

中文翻译：

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带有四个纳米抗体的 Fc 二聚体的生物化学、结构和细胞内化

VHH 仅代表骆驼 IgG 的重链可变区。VHH 家族形成了一组有趣的蛋白质，这些蛋白质来源于抗体，尽管它们的分子量相对较小（在 12,000 Da 范围内），但仍保持其识别抗原的能力。继续探索这些分子的可能性，我们选择通过将四个 VHH 与一个 Fc（抗体的片段可结晶区域）融合来设计具有保持抗原识别但增强能力的替代分子。在此过程中，我们的目标是获得具有出色定量抗原识别 (×4) 的分子，同时将其大小保持在 110 kDa 以下。在本文中，我们描述了我们创造的那些分子的构建 VHH ₂ -Fc-VHH ₂。VHH的结构₂ -Fc-VHH ₂与 HER2 抗原的复合物使用电子显微镜和建模来确定。该分子显示在其柔性臂末端结合四种 HER2 蛋白。VHH ₂ -Fc-VHH ₂还显示出优于参考抗 HER2 单克隆抗体 Herceptin® 的通过 HER2 受体的内化能力，并且优于两个 VHH 与一个 Fc (VHH ₂ -Fc) 的简单融合。这种新型分子，VHH ₂ -Fc-VHH ₂，可能是治疗库的有趣补充，具有从诊断到治疗的多种应用。