A multidisciplinary research program is designed to understand the structural basis of substrate specificity of the protein-tyrosine phosphates (PTPases) and to develop specific, high-affinity low molecular weight nonpeptidic PTPase inhibitors. PTPases constitute a large family of enzymes that can be collectively categorized into two groups: receptor-like and intracellular PTPases. The receptor-like PTPases, exemplified by PTP-alpha, generally have an extracellular domain, a single transmembrane region, and one or two cytoplasmic PTPase domains. The intracellular PTPases, exemplified by PTP1B, contain a single catalytic domain and various amino or carboxyl terminal extensions including SH2 domains that may have targeting or regulatory specificity phosphatases. VHR is a representative of the dual specificity phosphatases that can utilize substrates containing phosphotyrosine as well as phosphoserine/phosphothreonine. Although ligand binding and localization/targeting domains are believed to be involved in the regulation of PTPase activity, the role of the PTPase catalytic site in determining substrate specificity has not been fully investigated. This proposal will be focused on PTP1B, PTP-alpha and VHR. The substrate sequence specificity of these enzymes will be evaluated via combinatorial phosphopeptide libraries in combination with detailed kinetic analyses of individual pure peptides with systematically altered chemical properties. Active site-directed inhibitors will be identifies by screening synthetic libraries of low molecular weight aromatic phosphonates. The in vivo efficacy of selected inhibitors will be assessed in a Chinese hamster cell line that overexpresses human insulin receptor. Since the elucidation of the fundamentals of molecular recognition events requires an understanding of the atomic details of how molecules interact, the three-dimensional structures of PTPases complexed with high affinity substrates/inhibitors will be determined. PTP1B, PTP-alpha and VHR are ideal in this regard, since their crystal structures have been solved. Thus the crystal structures of these PTPases complexed with tight-binding substrates/ inhibitors can be solved by molecular replacement techniques.

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Research Project (R01)
Project #
5R01GM055242-03
Application #
6180654
Study Section
Biochemistry Study Section (BIO)
Program Officer
Ikeda, Richard A
Project Start
1998-09-30
Project End
2002-09-29
Budget Start
2000-09-30
Budget End
2001-09-29
Support Year
3
Fiscal Year
2000
Total Cost
$220,785
Indirect Cost
Name
Albert Einstein College of Medicine
Department
Pharmacology
Type
Schools of Medicine
DUNS #
009095365
City
Bronx
State
NY
Country
United States
Zip Code
10461
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Zhang, Zhong-Yin (2003) Mechanistic studies on protein tyrosine phosphatases. Prog Nucleic Acid Res Mol Biol 73:171-220
Sun, Jin-Peng; Fedorov, Alexander A; Lee, Seung-Yub et al. (2003) Crystal structure of PTP1B complexed with a potent and selective bidentate inhibitor. J Biol Chem 278:12406-14
Zhang, Zhong-Yin (2002) Protein tyrosine phosphatases: structure and function, substrate specificity, and inhibitor development. Annu Rev Pharmacol Toxicol 42:209-34
Zhang, Zhong-Yin; Zhou, Bo; Xie, Laiping (2002) Modulation of protein kinase signaling by protein phosphatases and inhibitors. Pharmacol Ther 93:307-17
Xie, Laiping; Zhang, Yan-Ling; Zhang, Zhong-Yin (2002) Design and characterization of an improved protein tyrosine phosphatase substrate-trapping mutant. Biochemistry 41:4032-9
Guo, Xiao-Ling; Shen, Kui; Wang, Fang et al. (2002) Probing the molecular basis for potent and selective protein-tyrosine phosphatase 1B inhibition. J Biol Chem 277:41014-22
Zhou, B; Wu, L; Shen, K et al. (2001) Multiple regions of MAP kinase phosphatase 3 are involved in its recognition and activation by ERK2. J Biol Chem 276:6506-15
Gao, Y; Voigt, J; Zhao, H et al. (2001) Utilization of a peptide lead for the discovery of a novel PTP1B-binding motif. J Med Chem 44:2869-78

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