Department of Chemistry, University of Zurich, Zurich, Switzerland.
Department of Chemical Sciences, Tata Institute of Fundamental Research, Mumbai, India.
Nat Protoc. 2020 Nov;15(11):3579-3594. doi: 10.1038/s41596-020-0386-5. Epub 2020 Oct 7.
Efficient methods to functionalize proteins are essential for the development of many diagnostic and therapeutic compounds, such as fluorescent probes for immunohistochemistry, zirconium-89 radiolabeled mAbs (Zr-mAbs) for positron emission tomography and antibody-drug conjugates (ADCs). This protocol describes a step-by-step procedure for the light-induced functionalization of proteins with compounds bearing the photochemically active aryl azide group. As an illustration of the potential utility of our approach, this protocol focuses on the synthesis of Zr-mAbs using photoactivatable derivatives of the metal ion binding chelate desferrioxamine B (DFO). The light-induced synthesis of Zr-mAbs is a unique, one-pot process involving simultaneous radiolabeling and protein conjugation. The photoradiochemical synthesis of purified Zr-mAbs, starting from unmodified proteins, [Zr][Zr(CO)] (Zr-oxalate), and a photoactivatable DFO derivative, can be performed in <90 min. The method can be easily adapted to prepare other radiolabeled proteins, ADCs or fluorescently tagged proteins by using drug molecules or fluorophores functionalized with photoactive moieties.
高效的蛋白质功能化方法对于许多诊断和治疗化合物的开发至关重要，例如用于免疫组织化学的荧光探针、用于正电子发射断层扫描的锆-89 放射性标记单克隆抗体 (Zr-mAb) 和抗体药物偶联物 (ADC)。本方案描述了一种逐步程序，用于用光诱导方法将具有光活性芳基叠氮基团的化合物功能化蛋白质。作为我们方法潜在用途的说明，本方案专注于使用金属离子结合螯合剂去铁胺 B (DFO) 的光活化衍生物合成 Zr-mAb。Zr-mAb 的光诱导合成是一种独特的、一锅法过程，涉及同时放射性标记和蛋白质偶联。从未修饰的蛋白质、[Zr][Zr(CO)] (Zr-草酸盐) 和光活化 DFO 衍生物开始，可在 <90 分钟内完成纯化 Zr-mAb 的光放射化学合成。该方法可以通过使用带有光活性部分的药物分子或荧光团很容易地适应制备其他放射性标记的蛋白质、ADC 或荧光标记的蛋白质。
