Gao Liqian, Sun Hongyan, Yao Shao Q
Department of Chemistry, Program of the Life Sciences Institute, National University of Singapore, 3 Science Drive 3, Singapore 117543, Singapore.
Biopolymers. 2010;94(6):810-9. doi: 10.1002/bip.21533.
Protein tyrosine phosphatases (PTPs) constitute a large family of enzymes that play key roles in cell signaling. Malfunctions of PTP activity have been linked to major human diseases including cancer. One key aspect in PTP biology is the elucidation of roles of PTPs, as well as substrates they act on, in different cellular events. Herein, a library of 144 putative peptide substrates against different PTPs was synthesized and immobilized onto a glass slide to generate the corresponding phosphopeptide microarray. Subsequent screening of the microarray against various PTPs provided a distinctive and comparative substrate fingerprint against each PTP. Several new substrates were identified, which might aid in the future design of potent and selective PTPs inhibitors. The signal-decrease microarray assay used in our studies provided a facile and efficient way for high-throughput determination of kinetic constants for peptide/PTP interactions en masse. Finally, our microarray results were independently verified by traditional microplate-based enzymatic assays.
蛋白质酪氨酸磷酸酶（PTPs）构成了一个在细胞信号传导中起关键作用的酶大家族。PTP活性的功能失调与包括癌症在内的主要人类疾病有关。PTP生物学的一个关键方面是阐明PTPs及其作用底物在不同细胞事件中的作用。在此，合成了一个针对不同PTPs的144种假定肽底物文库，并将其固定在载玻片上，以生成相应的磷酸肽微阵列。随后针对各种PTPs对微阵列进行筛选，得到了每种PTP独特的比较底物指纹图谱。鉴定出了几种新底物，这可能有助于未来设计强效和选择性的PTPs抑制剂。我们研究中使用的信号降低微阵列分析为高通量批量测定肽/PTP相互作用的动力学常数提供了一种简便有效的方法。最后，我们的微阵列结果通过传统的基于微孔板的酶促分析进行了独立验证。
