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Inactivation of rabbit muscle glycogen phosphorylase b by peroxynitrite revisited: does the nitration of Tyr613 in the allosteric inhibition site control enzymatic function?
Sharov, Victor S. Galeva, Nadezhda A. Dremina, Elena S. Williams, Todd D. Schoneich, Christian
Sharov, Victor S.
Galeva, Nadezhda A.
Dremina, Elena S.
Williams, Todd D.
Schoneich, Christian
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Abstract
There is increasing evidence that sequence-specific formation of 3-nitrotyrosine (3-NT) may cause functional changes in target proteins. Recently, the nitration of Tyr residues in glycogen phosphorylase b (Ph-b) was implicated in the age-associated decline of protein function (Sharov et al., Exp. Gerontol. 41, 407–416; 2006); in another report, the nitration of one specific residue, Tyr613, located in the allosteric inhibition site was hypothesized as a rationale for peroxynitrite inactivation (Dairou et al., J. Mol. Biol. 372, 1009–1021; 2007). In the present study, we have optimized the analysis of in-gel Ph-b digests by high performance liquid chromatography-electro spray ionization-tandem mass spectrometry, in order to achieve a quantitative analysis of nitration of individual Tyr residues at a high coverage of Tyr-containing sequences (92%). Our data do not confirm the role of Tyr613 nitration in the control of enzymatic function. Furthermore, we show here that the enzymatic activity of Ph-b does not directly correlate with the protein nitration levels, and that the modification of Cys and, potentially, other amino acid residues can better rationalize Ph-b inactivation by peroxynitrite.
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Date
2008-12-27
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Elsevier
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Keywords
Glycogen phosphorylase b, Enzymatic activity, Tryosine nitration, Cysteine oxidation, Peroxynitrite, Mass spectrometry, Solvent accessible surface area
Citation
Sharov, Victor S., Nadezhda A. Galeva, Elena S. Dremina, Todd D. Williams, and Christian Schöneich. "Inactivation of Rabbit Muscle Glycogen Phosphorylase B by Peroxynitrite Revisited: Does the Nitration of Tyr613 in the Allosteric Inhibition Site Control Enzymatic Function?" Archives of Biochemistry and Biophysics 484.2 (2009): 155-66.