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Activity‐based high‐throughput determination of PTPs substrate specificity using a phosphopeptide microarray
Author(s) -
Gao Liqian,
Sun Hongyan,
Yao Shao Q.
Publication year - 2010
Publication title -
peptide science
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.556
H-Index - 125
eISSN - 1097-0282
pISSN - 0006-3525
DOI - 10.1002/bip.21533
Subject(s) - phosphopeptide , chemistry , microarray , high throughput screening , enzyme , protein tyrosine phosphatase , peptide , substrate (aquarium) , protein microarray , microarray analysis techniques , computational biology , biochemistry , biology , gene expression , gene , ecology
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. © 2010 Wiley Periodicals, Inc. Biopolymers (Pept Sci) 94: 810–819, 2010. This article was originally published online as an accepted preprint. The “Published Online” date corresponds to the preprint version. You can request a copy of the preprint by emailing the Biopolymers editorial office at biopolymers@wiley.com