Site-Specific Encoding of Photoactivity in Antibodies Enables Light-Mediated Antibody-Antigen Binding on Live Cells
Site-Specific Encoding of Photoactivity in Antibodies Enables Light-Mediated Antibody-Antigen Binding on Live Cells
复制标题
抗体中光活性的位点特异性编码使得光介导的抗体-抗原能够在活细胞上结合
DOI：
10.1002/ange.201908655
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发表时间：
2019
影响因子：
--
通讯作者：
Bridge T
中科院分区：
文献类型：
--
作者：
Bridge T
抗体已在包括医学、诊断学和纳米技术的若干领域中得到应用，但使用外部试剂调节抗体-抗原结合的方法仍然有限。在这里，我们通过在抗体片段7 D12中用光笼化酪氨酸对单个酪氨酸残基进行遗传位点特异性置换，开发了光活性抗体片段。采用一种简单而稳健的测定法来评估7 D12突变体与癌细胞表面上其靶点表皮生长因子受体（EGFR）的光介导结合。在所研究的三种7 D12突变体中的两种中，光笼化酪氨酸的存在使7 D12-EGFR结合亲和力降低了20倍以上，并且在暴露于365 nm光时恢复结合。 分子动力学模拟解释了突变体之间光笼化对7 D12-EGFR相互作用的影响的差异。最后，我们展示了光敏抗体在以光依赖性方式将荧光团递送至EGFR阳性活癌细胞中的应用。
Antibodies have found applications in several fields, including, medicine, diagnostics, and nanotechnology, yet methods to modulate antibody–antigen binding using an external agent remain limited. Here, we have developed photoactive antibody fragments by genetic site‐specific replacement of single tyrosine residues with photocaged tyrosine, in an antibody fragment, 7D12. A simple and robust assay is adopted to evaluate the light‐mediated binding of 7D12 mutants to its target, epidermal growth factor receptor (EGFR), on the surface of cancer cells. Presence of photocaged tyrosine reduces 7D12‐EGFR binding affinity by over 20‐fold in two out of three 7D12 mutants studied, and binding is restored upon exposure to 365 nm light. Molecular dynamics simulations explain the difference in effect of photocaging on 7D12‐EGFR interaction among the mutants. Finally, we demonstrate the application of photoactive antibodies in delivering fluorophores to EGFR‐positive live cancer cells in a light‐dependent manner.