Jensen Rasmus K, Jabs Frederic, Miehe Michaela, Mølgaard Brian, Pfützner Wolfgang, Möbs Christian, Spillner Edzard, Andersen Gregers R
Department of Molecular Biology and Genetics, Aarhus University, Aarhus, Denmark.
Immunological Biotechnology, Department of Engineering, Aarhus University, Aarhus, Denmark.
Allergy. 2020 Aug;75(8):1956-1965. doi: 10.1111/all.14222. Epub 2020 Mar 9.
IgE is the central antibody isotype in TH2-biased immunity and allergic diseases. The structure of intact IgE and the impact of IgE-targeting molecules on IgE however remain elusive. In order to obtain insights into IgE biology and the clinical impact, we aimed for structure determination of IgE and the complex of IgE with the anti-IgE antibody ligelizumab.
Structures of two distinct intact IgE with specificity for cross-reactive carbohydrate determinants and Der p 2 as well as complexes of ligelizumab-Fab with IgE and IgE Fc were assessed by negative stain electron microscopy and solution scattering. Inhibition of IgE binding and displacement of receptor-bound IgE were assessed using cellular assays, basophil activation testing and ELIFAB assays.
Our data reveal that the investigated IgE molecules share an overall rigid conformation. In contrast to the IgE Fc fragment, the IgE Fc in intact IgE is significantly less asymmetrically bent. The proximal and the distal Fabs are rigidly tethered to the Fc. Binding of ligelizumab to IgE in a 2:1 stoichiometry induces an extended and twofold symmetrical conformation of IgE, which retains a rigid Fab-Fc architecture. Analyses of effector cell activation revealed that ligelizumab inhibits IgE binding without displacing receptor-bound IgE. Together with an interference of CD23 binding, the data underline a functional activity similar to omalizumab.
Our data reveal the first structures of intact IgE suggesting that the IgE Fab is fixed relative to the Fc. Furthermore, we provide a structural rationale for the inhibitory mechanism of ligelizumab.
IgE是2型辅助性T细胞（TH2）偏向性免疫和过敏性疾病中的核心抗体同种型。然而，完整IgE的结构以及靶向IgE分子对IgE的影响仍不清楚。为了深入了解IgE生物学特性及其临床影响，我们旨在确定IgE以及IgE与抗IgE抗体利吉珠单抗复合物的结构。
通过负染电子显微镜和溶液散射评估了两种对交叉反应性碳水化合物决定簇和Der p 2具有特异性的不同完整IgE的结构，以及利吉珠单抗Fab与IgE和IgE Fc的复合物。使用细胞试验、嗜碱性粒细胞激活试验和ELIFAB试验评估IgE结合的抑制和受体结合IgE的置换。
我们的数据表明，所研究的IgE分子具有整体刚性构象。与IgE Fc片段相比，完整IgE中的IgE Fc不对称弯曲程度明显较小。近端和远端Fab牢固地连接到Fc上。利吉珠单抗以2:1的化学计量比与IgE结合会诱导IgE形成伸展且具有双重对称性的构象，该构象保留了刚性的Fab-Fc结构。效应细胞激活分析表明，利吉珠单抗抑制IgE结合但不会置换受体结合的IgE。连同对CD23结合的干扰，这些数据强调了其与奥马珠单抗相似的功能活性。
我们的数据揭示了完整IgE的首个结构，表明IgE Fab相对于Fc是固定的。此外，我们为利吉珠单抗的抑制机制提供了结构依据。
