1pa1

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spinqualitylabelsall models 1pa1, resolution 1.6Å Show:Asymmetric Unit Biological Assembly Drag the structure with the mouse to rotate   Export Animated Image

Crystal structure of the C215D mutant of protein tyrosine phosphatase 1B

Contents 1 Overview 2 Disease 3 About this Structure 4 Reference

Overview

We have characterized the C215D active-site mutant of protein-tyrosine, phosphatase-1B (PTP-1B) and solved the crystal structure of the catalytic, domain of the apoenzyme to a resolution of 1.6 A. The mutant enzyme, displayed maximal catalytic activity at pH approximately 4.5, which is, significantly lower than the pH optimum of 6 for wild-type PTP-1B., Although both forms of the enzyme exhibited identical Km values for, hydrolysis of p-nitrophenyl phosphate at pH 4.5 and 6, the kcat values of, C215D were approximately 70- and approximately 7000-fold lower than those, of wild-type PTP-1B, respectively. Arrhenius plots revealed that the, mutant and wild-type enzymes displayed activation energies of 61 +/- 1 and, 18 +/- 2 kJ/mol, respectively, at their pH optima. Unlike wild-type, PTP-1B, C215D-mediated p-nitrophenyl phosphate hydrolysis was inactivated, by 1,2-epoxy-3-(p-nitrophenoxy)propane, suggesting a direct involvement of, Asp215 in catalysis. Increasing solvent microviscosity with sucrose (up to, 40% (w/v)) caused a significant decrease in kcat/Km of the wild-type, enzyme, but did not alter the catalytic efficiency of the mutant protein., Structurally, the apoenzyme was identical to wild-type PTP-1B, aside from, the flexible WPD loop region, which was in both "open" and "closed", conformations. At physiological pH, the C215D mutant of PTP-1B should be, an effective substrate-trapping mutant that can be used to identify, cellular substrates of PTP-1B. In addition, because of its insensitivity, to oxidation, this mutant may be used for screening fermentation broth and, other natural products to identify inhibitors of PTP-1B.

Disease

Known diseases associated with this structure: Abdominal body fat distribution, modifier of OMIM:[176885], Insulin resistance, susceptibility to OMIM:[176885]

About this Structure

1PA1 is a Single protein structure of sequence from Homo sapiens with MG and CL as ligands. Active as Protein-tyrosine-phosphatase, with EC number 3.1.3.48 Full crystallographic information is available from OCA.

Reference

Functional characterization and crystal structure of the C215D mutant of protein-tyrosine phosphatase-1B., Romsicki Y, Scapin G, Beaulieu-Audy V, Patel S, Becker JW, Kennedy BP, Asante-Appiah E, J Biol Chem. 2003 Aug 1;278(31):29009-15. Epub 2003 May 13. PMID:12748196

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