Department of Biomedical Sciences, Graduate School, Ajou University, 206 World cup-ro, Yeongtong-gu, Suwon, 16499, Gyeonggi-do, South Korea.
Department of Microbiology, Ajou University School of Medicine, 206 World cup-ro, Yeongtong-gu, Suwon, 16499, Gyeonggi-do, South Korea.
Sci Rep. 2020 Feb 7;10(1):2140. doi: 10.1038/s41598-020-58798-7.
Intrabodies, antibodies expressed within cells, offer an interesting way to target intracellular molecules, making them potentially useful for biotechnology and medicine. However, it remains controversial whether full-size IgG intrabodies expressed in the reducing environment of the cytosol of mammalian cells are workable and structurally sound. Herein, we settle this issue with a systematic investigation of the structure and functionality of four chimeric IgG1s with distinct variable (V) domains but identical constant (C) domains. Full-size IgGs expressed in the cytosol of HEK293 cells were either assembly-competent or -incompetent, depending on the intrinsic properties of the V regions. Structural integrity of the C region is required for H:L association and the formation of a functional antigen-binding site. Partial intrachain disulfide bond formation occurs in both H and L chains of cytosolic IgG intrabodies, whereas interchain disulfide bond formation was absent and dispensable for functional assembly. IgG1s expressed in the cytosol and via the ER were shown to assemble differently. Our findings provide insight into the features and possible utilization of full-size IgGs as cytosolic antibodies in biotechnological and medical applications.
内抗体是在细胞内表达的抗体，为靶向细胞内分子提供了一种有趣的方法，因此它们在生物技术和医学领域具有潜在的应用价值。然而，在哺乳动物细胞胞质的还原环境中表达的完整 IgG 内抗体是否可行且结构合理，这一问题仍存在争议。本文通过对具有不同可变 (V) 区但相同恒定 (C) 区的四个嵌合 IgG1 的结构和功能进行系统研究，解决了这一问题。在 HEK293 细胞的胞质中表达的全长 IgG 要么具有组装能力，要么不具有组装能力，这取决于 V 区的固有特性。C 区的结构完整性对于 H:L 结合和形成功能性抗原结合位点是必需的。胞质内 IgG 内抗体的 H 链和 L 链中均会发生部分链内二硫键形成，而链间二硫键形成则不存在且对于功能性组装是可有可无的。在胞质中以及通过内质网表达的 IgG1 被证明具有不同的组装方式。我们的研究结果为全面了解作为生物技术和医学应用中胞质抗体的完整 IgG 的特性和可能用途提供了线索。
