Complementary Structural Information for Antibody-Antigen Complexes from Hydrogen-Deuterium Exchange and Covalent Labeling Mass Spectrometry.
Complementary Structural Information for Antibody-Antigen Complexes from Hydrogen-Deuterium Exchange and Covalent Labeling Mass Spectrometry.
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DOI：
10.1021/jasms.2c00108
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发表时间：
2022-07-06
影响因子：
3.2
通讯作者：
Vachet, Richard W.
中科院分区：
文献类型：
--
作者：
Tremblay, Catherine Y.;Kirsch, Zachary J.;Vachet, Richard W.
表征抗体-抗原相互作用对于正确开发治疗性抗体、理解其作用机制以及为新药分子申请专利是必要的。在这里，我们证明了氢氘交换（HDX）质谱（MS）测量与焦碳酸二乙酯（DEPC）共价标记（CL）MS测量一起提供了关于抗体-抗原相互作用的高阶结构信息，这是单独使用任何一种技术都无法获得的。使用肿瘤坏死因子α（TNFα）与三种不同单克隆抗体（mAb）复合的良好表征的模型系统，我们表明两种技术提供了结合不同mAb后TNFα结构变化的更完整的整体图像，有时提供了有关结合位点和结合后蛋白质动力学变化的协同信息。CL的标记降低通常发生在TNFα表位附近，而HDX的降低可能跨越整个蛋白质，因为当mAb结合TNFα时会发生实质性稳定。综合考虑两个数据集，阐明了因mAb结合而导致溶剂暴露量降低和因mAb结合而导致动力学变化的TNFα区域。此外，DEPC-CL/MS的单残基水平分辨率可以澄清长肽的HDX/MS数据。我们认为，这两种技术应该一起使用时，研究单克隆抗体-抗原相互作用，因为它们提供的互补信息。基于焦碳酸二乙酯的共价标记和氢-氘交换质谱法提供了关于抗体-抗原结合相互作用的互补和协同的结构信息。
Characterizing antibody-antigen interactions is necessary for properly developing therapeutic antibodies, understanding their mechanisms of action, and patenting new drug molecules. Here, we demonstrate that hydrogen deuterium exchange (HDX) mass spectrometry (MS) measurements together with diethylpyrocarbonate (DEPC) covalent labeling (CL) MS measurements provides higher-order structural information about antibody-antigen interactions that is not available from either technique alone. Using the well-characterized model system of tumor necrosis factor α (TNFα) in complex with three different monoclonal antibodies (mAbs), we show that two techniques offer a more complete overall picture of TNFα’s structural changes upon binding different mAbs, sometimes providing synergistic information about binding sites and changes in protein dynamics upon binding. Labeling decreases in CL generally occur near the TNFα epitope, whereas decreases in HDX can span the entire protein due to substantial stabilization that occurs when mAbs bind TNFα. Considering both datasets together clarifies the TNFα regions that undergo a decrease in solvent exposure due to mAb binding and that undergo a change in dynamics due to mAb binding. Moreover, the single-residue level resolution of DEPC-CL/MS can clarify HDX/MS data for long peptides. We feel that the two techniques should be used together when studying the mAb-antigen interactions because of the complementary information they provide. Diethylpyrocarbonate-based covalent labeling and hydrogen-deuterium exchange mass spectrometry provide complementary and synergistic structural information about antibody-antigen binding interactions.
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DOI：
10.1021/jasms.1c00373
发表时间：
2022-03-02
影响因子：
3.2
作者：
Biehn SE;Picarello DM;Pan X;Vachet RW;Lindert S
通讯作者：
Lindert S
影响因子：
48
作者：
Masson, Glenn R.;Burke, John E.;Rand, Kasper D.
通讯作者：
Rand, Kasper D.
影响因子：
4.3
作者：
Huang，Richard Y. -C.;Kuhne，Michelle;Chen，Guodong
通讯作者：
Chen，Guodong
DOI：
10.1007/s13361-017-1815-8
发表时间：
2018-01-01
影响因子：
3.2
作者：
Huang, Richard Y. -C.;Iacob, Roxana E.;Chen, Guodong
通讯作者：
Chen, Guodong
影响因子：
7.4
作者：
Liu, Tianying;Limpikirati, Patanachai;Vachet, Richard W.
通讯作者：
Vachet, Richard W.