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Báo cáo khoa học: Effect of ionic strength and oxidation on the P-loop conformation of the protein tyrosine phosphatase-like phytase, PhyAsr

The protein tyrosine phosphatase (PTP)-like phytase, PhyAsr, fromSeleno-monas ruminantiumis a novel member of the PTP superfamily, and the only described member that hydrolyzes myo-inositol-1,2,3,4,5,6-hexakisphosphate. | ỊFEBS Journal Effect of ionic strength and oxidation on the P-loop conformation of the protein tyrosine phosphatase-like phytase PhyAsr Robert J. Gruninger1 L. Brent Selinger2 and Steven C. Mosimann1 1 Department of Chemistry and Biochemistry University of Lethbridge Canada 2 Department of BiologicalSciences University of Lethbridge Canada Keywords ionic strength oxidation phytase P-loop protein tyrosine phosphatase Correspondence S. C. Mosimann Department of Chemistry and Biochemistry University of Lethbridge Lethbridge AB Canada T1K 3M4 Fax 1 403 329 2057 Tel 1 403 329 2283 E-mail steven.mosimann@uleth.ca Database The coordinates and structure factors for the structures of PhyAsr at ionic strengths of 200 300 400 and 500 mM and with the catalytic cysteine oxidized to cysteine sulfonic acid have been deposited in the Protein Data Bank entries 2PSZ 3D1O 3D1Q 3D1H and 2PT0 respectively Received 26 March 2008 revised 21 May 2008 accepted 27 May 2008 doi 10.1111 j.1742-4658.2008.06524.x The protein tyrosine phosphatase PTP -like phytase PhyAsr from Seleno-monas ruminantium is a novel member of the PTP superfamily and the only described member that hydrolyzes myo-inositol-1 2 3 4 5 6-hexakisphosphate. In addition to the unique substrate specificity of PhyAsr the phosphate-binding loop P-loop has been reported to undergo a conformational change from an open inactive to a closed active conformation upon ligand binding at low ionic strength. At high ionic strengths the P-loop was observed in the closed active conformation in both the presence and absence of ligand. To test whether the P-loop movement can be induced by changes in ionic strength we examined the effect that ionic strength has on the catalytic efficiency of PhyAsr and determined the structure of the enzyme at several ionic strengths. The catalytic efficiency of PhyAsr is highly sensitive to ionic strength with a seven-fold increase in kcat Km and a ninefold decrease in Km when the ionic strength is increased