Ligand-Directed N-Sulfonyl Pyridone Chemistry for Selective Native Protein Labeling and Imaging in Live Cell
Ligand-Directed N-Sulfonyl Pyridone Chemistry for Selective Native Protein Labeling and Imaging in Live Cell
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DOI：
10.1007/978-1-4939-9537-0_16
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发表时间：
2019-01-01
期刊：
影响因子：
--
通讯作者：
Hamachi, Itaru
中科院分区：
文献类型：
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作者：
Masuda, Marie;Matsuo, Kazuya;Hamachi, Itaru
应用于活细胞条件下内源性蛋白质修饰的生物相容性有机化学的进展已经很长时间了，因为这些可以为阐明各种生物现象提供重要的工具。但是，目前还存在多种障碍，如反应模式的储备有限、反应动力学缓慢、内源性蛋白修饰的特异性不足等。我们最近报道了一种新的基于亲和的标记技术，称为配体定向（LD）化学，它不需要任何遗传操作，这与其他策略形成鲜明对比，包括肽/酶标记方法或基于生物正交化学的方法。在这里，我们描述了LD化学的一般原理，使用n -磺酰基吡啶酮（SP）作为一种新的反应基团（LDSP化学），它允许内源性蛋白质磺化，具有更高的标记率和特异性，相对于我们之前报道的活细胞表面和内部的LD化学。详细的协议LDSP化学碳酸酐酶标记和成像在体外和活细胞解释。
Advances in biocompatible organic chemistry applicable for endogenous protein modification under live-cell conditions have been longed as these can produce an important tool for the elucidation of a variety of biological phenomena. However, there are still various obstacles to be overcome, such as the limited repertories of the reaction modes, the slow reaction kinetics, and the insufficient specificity for endogenous protein modification. We have recently reported a new type of affinity-based labeling technique termed ligand-directed (LD) chemistry that does not need any genetic manipulation, which shows a sharp contrast with other strategies including peptide/enzyme-tag methods or bioorthogonal chemistry-based methods. Here we describe the general principles of LD chemistry using N-sulfonyl pyridone (SP) as a new reactive group (LDSP chemistry) that allows for endogenous protein sulfonylation with the higher labeling rate and specificity, relative to our previously reported LD chemistry on the surface of and the inside of live cells. The detailed protocols of LDSP chemistry for carbonic anhydrase labeling and imaging in vitro and in living cells are explained.