Polyproline-rod approach to isolating protein targets of bioactive small molecules: Isolation of a new target of indomethacin
Polyproline-rod approach to isolating protein targets of bioactive small molecules: Isolation of a new target of indomethacin
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DOI：
10.1021/ja0655643
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发表时间：
2007-01-31
影响因子：
15
通讯作者：
Uesugi, Motonari
中科院分区：
文献类型：
--
作者：
Sato, Shin-ichi;Kwon, Youngjoo;Uesugi, Motonari
生物活性小分子靶蛋白的鉴定一直是化学遗传学的技术难点。在这里，我们报告了一个聚脯氨酸棒的方法，从细胞裂解液中分离小分子的蛋白质靶点。结果表明，插入一个长的，刚性的聚脯氨酸螺旋之间的小分子诱饵和生物素标签提高亲和纯化的能力，从而允许低丰度或低亲和力的蛋白质的分离。在概念验证实验的过程中，我们分离出了作为吲哚美辛（一种广泛使用的消炎药）新靶点的谷胱甘肽酶1（GLO1）。分子生物学实验表明，GLO1酶活性的抑制与临床公认的吲哚美辛家族非甾体抗炎药的有益副作用有关。
Identification of protein targets of bioactive small molecules has been a technical hurdle of chemical genetics. Here we report a polyproline-rod approach to isolating protein targets of small molecules from cell lysates. The results indicate that insertion of a long, rigid polyproline helix between a small-molecule bait and a biotin tag boosts the capacity of affinity purification and thereby permits isolation of low-abundance or low-affinity proteins. In the course of the proof-of-concept experiments, we isolated glyoxalase 1 (GLO1) as a new target of indomethacin, a widely used antiinflammatory drug. Molecular biological experiments suggest that inhibition of GLO1 enzyme activity is related to the clinically recognized beneficial side effects of the indomethacin family of nonsteroidal antiinflammatory drugs.