Metal ion-mediated polymer superquenching for highly sensitive detection of kinase and phosphatase activities

被引：125

作者：

Rininsland, F ^{[1
]}

Xia, WS ^{[1
]}

Wittenburg, S ^{[1
]}

Shi, XB ^{[1
]}

Stankewicz, C ^{[1
]}

Achyuthan, K ^{[1
]}

McBranch, D ^{[1
]}

Whitten, D ^{[1
]}

机构：

[1] QTL Biosyst, Santa Fe, NM 87507 USA

来源：

PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA | 2004年 / 101卷 / 43期

关键词：

D O I：

10.1073/pnas.0406832101

中图分类号：

O [数理科学和化学]; P [天文学、地球科学]; Q [生物科学]; N [自然科学总论];

学科分类号：

07 ; 0710 ; 09 ;

摘要：

An assay technology for high-throughput screening of kinase and phosphatase activities is introduced. The format is based upon superquenching of fluorescent-conjugated polymers by dye-labeled kinase/phosphatase peptide substrates. The sensor platform is composed of highly fluorescent-conjugated polyelectrolytes colocated with the phosphate coordinating metal ion gallium on microspheres. Phosphorylated peptide substrates containing a quencher bind specifically to the metal ions by means of phosphate groups, resulting in quench of polymer fluorescence. The modulation of fluorescence signal is proportional to kinase or phosphatase activity and is monitored as a turn-off or turn-on signal, respectively. The assay is homogeneous and simple and can be run either as an endpoint measurement or in a kinetic mode. The assay meets the sensitivity required for high-throughput screening of kinase or phosphatase inhibitors and is a valuable tool for drug discovery. A modified version of the assay allows for the detection of protein phosphorylation.

引用

页码：15295 / 15300

页数：6

共 40 条

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