The protein tyrosine phosphatases (PTPs), which dephosphorylate specific phosphotyrosine residues in protein substrates, constitute a large family of signaling enzymes, whose activity is ubiquitously misregulated in human disease. Specifically, aberrant PTP activity has been implicated as a causative agent in leukemias, solid-tumor cancers, type I and type II diabetes, and a host of autoimmune disorders. Tools that are capable of inducing target- specific PTP inhibition or activation would be invaluable for delineation of the precise functions of PTPs in cell signaling and for the validation of PTPs as therapeutic targets. However, the common architecture of the conserved PTP protein fold impedes the discovery of selective PTP inhibitors, and cell-permeable PTP activators have not been identified to date. The broad objective of the proposed research is to develop tools to specifically inhibit and activate engineered PTPs. To avoid the specificity problems that are inherent to conventional medicinal chemistry, we are using protein-engineering strategies for generating inhibitor- sensitive and activator-sensitive (collectively, ligand-sensitive) PTPs: engineered phosphatases whose activities are uniquely responsive to cell-permeable biarsenical small molecules. The crux of our protein-engineering strategies resides in the introduction of biarsenical-binding cysteine-rich motifs that are not present in wild-type PTPs. The engineered motifs are rationally designed to induce either PTP inhibition or activation upon binding of the biarsenical ligands, which do not affect the activities of naturally occurring PTPs. In a cellular context therefore, target-specific chemical control over the activity of a single PTP of interest can be affected by adding the ligand to cells that express the sensitized PTP. Our proposed experiments include optimization of a previously discovered strategy for sensitizing PTPs to inhibition by biarsenicals (Specific Aim 1: Biochemical optimization of PTP inhibition by targeting the WPD-loop);investigating the scope of application of a recently discovered strategy for chemically activating PTPs from an immunologically important sub- family (Specific Aim 2: Target-specific activation of the NT4 family of PTPs);inhibiting PTPs by targeting an allosteric site that is 20? removed from the conserved active site (Specific Aim 3: Targeting a PTP allosteric-inhibition site with biarsenical small molecules);and using target- specific inhibitors and activators to elucidate the signaling roles of an oncogenic PTP in living cells (Specific Aim 4: Target-specific inhibition and activation of the oncogenic PTP Shp2).

Public Health Relevance

Protein tyrosine phosphatases (PTPs) are enzymes that help to send cellular messages by enzymatically removing phosphate groups from other proteins. When cellular phosphate removal goes awry, so do the basic regulatory mechanisms of the cell, and improperly regulated PTP activity has been implicated as a causative agent in a range of human diseases, including cancer, diabetes, and autoimmune disorders. We are designing small-molecule PTP inhibitors and activators that can be used to study the functions of individual PTPs in cellular experiments;this work will thus provide valuable tools for dissecting cellular phosphorylation networks and for validating PTPs as potential therapeutic targets.

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Academic Research Enhancement Awards (AREA) (R15)
Project #
2R15GM071388-03
Application #
8367318
Study Section
Macromolecular Structure and Function E Study Section (MSFE)
Program Officer
Gerratana, Barbara
Project Start
2005-08-15
Project End
2015-07-31
Budget Start
2012-08-01
Budget End
2015-07-31
Support Year
3
Fiscal Year
2012
Total Cost
$292,391
Indirect Cost
$82,391
Name
Amherst College
Department
Chemistry
Type
Schools of Arts and Sciences
DUNS #
066985367
City
Amherst
State
MA
Country
United States
Zip Code
01002
Harris, Leigh K; Frumm, Stacey M; Bishop, Anthony C (2013) A general assay for monitoring the activities of protein tyrosine phosphatases in living eukaryotic cells. Anal Biochem 435:99-105
Davis, Oliver B; Bishop, Anthony C (2012) Specific inhibition of sensitized protein tyrosine phosphatase 1B (PTP1B) with a biarsenical probe. Bioconjug Chem 23:272-8
Walton, Zandra E; Bishop, Anthony C (2010) Target-specific control of lymphoid-specific protein tyrosine phosphatase (Lyp) activity. Bioorg Med Chem 18:4884-91
Chen, Vincent L; Bishop, Anthony C (2010) Chemical rescue of protein tyrosine phosphatase activity. Chem Commun (Camb) 46:637-9
Zhang, Xin-Yu; Chen, Vincent L; Rosen, Mari S et al. (2008) Allele-specific inhibition of divergent protein tyrosine phosphatases with a single small molecule. Bioorg Med Chem 16:8090-7
Zhang, Xin-Yu; Bishop, Anthony C (2008) Engineered inhibitor sensitivity in the WPD loop of a protein tyrosine phosphatase. Biochemistry 47:4491-500
Zhang, Xin-Yu; Bishop, Anthony C (2007) Site-specific incorporation of allosteric-inhibition sites in a protein tyrosine phosphatase. J Am Chem Soc 129:3812-3
Bishop, Anthony C; Zhang, Xin-Yu; Lone, Anna Mari (2007) Generation of inhibitor-sensitive protein tyrosine phosphatases via active-site mutations. Methods 42:278-88
Bishop, Anthony C; Blair, Elizabeth R (2006) A gatekeeper residue for inhibitor sensitization of protein tyrosine phosphatases. Bioorg Med Chem Lett 16:4002-6