Genetically targeted chromophore-assisted light inactivation
Genetically targeted chromophore-assisted light inactivation
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DOI：
10.1038/nbt914
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发表时间：
2003-12-01
影响因子：
46.9
通讯作者：
Tsien, RY
中科院分区：
文献类型：
--
作者：
Tour, O;Meijer, RM;Tsien, RY
蛋白质功能的研究将促进一个通用的方法，以非侵入性的高空间和时间精度的活细胞中的选定的蛋白质。发色团辅助光失活（卡利）（1）利用光化学产生的活性氧来快速灭活蛋白质，但其使用受到需要显微注射染料标记的非功能阻断抗体的限制。我们现在证明了连接蛋白43（Cx43）和α（1C）L型钙通道的卡利，每个通道都标记有一个或两个小的四半胱氨酸（TC）基序（2），这些基序特异性结合膜渗透性红色双砷染料ReAsH 3，4.基于ReAsH的卡利是基因靶向的，不需要抗体或显微注射，并且在体内灭活每种蛋白质的比例接近90%。
Studies of protein function would be facilitated by a general method to inactivate selected proteins in living cells noninvasively with high spatial and temporal precision. Chromophore- assisted light inactivation (CALI)(1) uses photochemically generated, reactive oxygen species to inactivate proteins acutely, but its use has been limited by the need to microinject dye- labeled nonfunction- blocking antibodies. We now demonstrate CALI of connexin43 (Cx43) and alpha(1C) L- type calcium channels, each tagged with one or two small tetracysteine (TC) motifs(2) that specifically bind the membrane- permeant, red biarsenical dye, ReAsH3,4. ReAsH-based CALI is genetically targeted, requires no antibodies or microinjection, and inactivates each protein by similar to 90% in