Higher-order structure (HOS) characterization of therapeutic protein-drug conjugates for comprehensive assessment of conjugation-induced protein conformational changes is an important consideration in the biopharmaceutical industry to ensure proper behavior of protein therapeutics. In this study, conformational dynamics of a small therapeutic protein, adnectin 1, together with its drug conjugate were characterized by hydrogen/deuterium exchange mass spectrometry (HDX-MS) with different spatial resolutions. Top-down HDX allows detailed assessment of the residue-level deuterium content in the payload conjugation region. HDX-MS dataset revealed the ability of peptide-based payload/linker to retain deuterium in HDX experiments. Combined results from intact, top-down, and bottom-up HDX indicated no significant conformational changes of adnectin 1 upon payload conjugation.

中文翻译：

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氢/氘交换质谱法对Adnectin和Adnectin-药物结合物的构象评估

治疗性蛋白质-药物偶联物的高阶结构（HOS）表征，用于综合评估偶联诱导的蛋白质构象变化，是生物制药行业中确保蛋白质治疗剂正确行为的重要考虑因素。在这项研究中，通过氢/氘交换质谱（HDX-MS）表征了具有不同空间分辨率的小型治疗性蛋白adnectin 1及其药物结合物的构象动力学。自上而下的HDX可以详细评估有效负载共轭区域中残留物水平的氘含量。HDX-MS数据集揭示了在HDX实验中基于肽的有效载荷/接头保留氘的能力。完整，自上而下，