Self-reporting fluorescent substrates of protein tyrosine kinases
Self-reporting fluorescent substrates of protein tyrosine kinases
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DOI：
10.1021/ja0577692
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发表时间：
2006-02-15
影响因子：
15
通讯作者：
Lawrence, DS
中科院分区：
文献类型：
--
作者：
Wang, QZ;Cahill, SM;Lawrence, DS
一个新的机制的原则，蛋白酪氨酸激酶底物荧光报告的磷酸基团的引入已经开发。NMR用于确定酪氨酸磷酸化诱导酪氨酸部分与肽底物上的近端荧光团的π−π堆积相互作用的破坏。我们已经证明，（1）本研究中描述的肽底物可用于各种不同的酪氨酸激酶，（2）可以采用生理浓度的ATP（与标准放射性ATP激酶测定不同），从而提供更真实的抑制剂效力评估，以及（3）可以实时观察蛋白激酶自激活。
A new mechanistic principle by which protein tyrosine kinase substrates fluorescently report the introduction of a phosphate moiety has been developed. NMR was used to establish that tyrosine phosphorylation induces the disruption of π−π stacking interactions of the tyrosine moiety with a proximal fluorophore on the peptide substrate. We have demonstrated that (1) the peptide substrates described in this study are useful for a wide variety of different tyrosine kinases, (2) physiological concentrations of ATP can be employed (unlike the standard radioactive ATP kinase assays), thus providing a more realistic assessment of inhibitor potency, and (3) protein kinase self-activation can be observed in real-time.