Effect of pH on the Oxidation-Reduction Properties of Thioredoxins

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• 作者：Aaron T. Setterdahl ; Peter T. Chivers ; Masakazu Hirasawa ; Sté ; phane D. Lemaire ; Eliane Keryer ; Myroslawa Miginiac-Maslow ; Sung-Kun Kim ; Jeremy Mason ; Jean-Pierre Jacquot ; Christopher C. Longbine
• 刊名：Biochemistry
• 出版年：2003
• 出版时间：December 23, 2003
• 年：2003
• 卷：42
• 期：50
• 页码：14877 - 14884
• 全文大小：91K
• 年卷期：v.42,no.50(December 23, 2003)
• ISSN：1520-4995

文摘

Oxidation-reduction midpoint potential (E_m) versus pH profiles were measured for wild-typethioredoxins from Escherichia coli and from the green alga Chlamydomonas reinhardtii and for a numberof site-directed mutants of these two thioredoxins. These profiles all exhibit slopes of approximately -59mV per pH unit, characteristic of the uptake of two protons per reduction of an active-site thioredoxindisulfide, at acidic, neutral, and moderately alkaline pH values. At higher pH values, these profiles exhibitslopes of either -29.5 mV per pH unit, characteristic of the uptake of one proton per disulfide reduced,or are pH-independent, indicating that neither proton uptake nor proton release is associated with reductionof the active-site disulfide. Reduction of the two wild-type thioredoxins is accompanied by the uptake oftwo protons even at pH values where the more acidic cysteine thiol group of the reduced proteins wouldbe expected to be completely unprotonated. The effect of site-directed mutagenesis of two highly conservedaspartate residues that play important structural and/or catalytic roles in both thioredoxins, and whichcould in principle play a role in proton transfer, on the pK_a values of redox-linked acid dissociations(deduced from changes in slope of the E_m versus pH profiles) has also been determined for both E. colithioredoxin and C. reinhardtii thioredoxin h.