Laboratoire de Biologie Moléculaire et Cellulaire des Eucaryotes, FRE3354 Centre National de la Recherche Scientifique, Institut de Biologie Physico-Chimique, Université Pierre et Marie Curies, Paris, France.
Antioxid Redox Signal. 2012 Jan 1;16(1):17-32. doi: 10.1089/ars.2011.3933. Epub 2011 Aug 30.
Cysteines (Cys) made acidic by the protein environment are generally sensitive to pro-oxidant molecules. Glutathionylation is a post-translational modification that can occur by spontaneous reaction of reduced glutathione (GSH) with oxidized Cys as sulfenic acids (-SOH). The reverse reaction (deglutathionylation) is strongly stimulated by glutaredoxins (Grx) and requires a reductant, often GSH.
Here, we show that chloroplast GrxS12 from poplar efficiently reacts with glutathionylated substrates in a GSH-dependent ping pong mechanism. The pK(a) of GrxS12 catalytic Cys is very low (3.9) and makes GrxS12 itself sensitive to oxidation by H(2)O(2) and to direct glutathionylation by nitrosoglutathione. Glutathionylated-GrxS12 (GrxS12-SSG) is temporarily inactive until it is deglutathionylated by GSH. The equilibrium between GrxS12 and glutathione (E(m(GrxS12-SSG))= -315 mV, pH 7.0) is characterized by K(ox) values of 310 at pH 7.0, as in darkened chloroplasts, and 69 at pH 7.9, as in illuminated chloroplasts.
Based on thermodynamic data, GrxS12-SSG is predicted to accumulate in vivo under conditions of mild oxidation of the GSH pool that may occur under stress. Moreover, GrxS12-SSG is predicted to be more stable in chloroplasts in the dark than in the light.
These peculiar catalytic and thermodynamic properties could allow GrxS12 to act as a stress-related redox sensor, thus allowing glutathione to play a signaling role through glutathionylation of GrxS12 target proteins.
在蛋白质环境中带负电荷的半胱氨酸通常对氧化剂分子敏感。谷胱甘肽化是一种翻译后修饰，它可以通过还原型谷胱甘肽（GSH）与氧化的半胱氨酸（Cys）反应生成亚磺酸（-SOH）自发发生。该反应的逆反应（去谷胱甘肽化）强烈地被谷氧还蛋白（Grx）刺激，并需要还原剂，通常是 GSH。
在这里，我们展示了杨树质体 GrxS12 能够以 GSH 依赖的乒乓机制有效地与谷胱甘肽化的底物反应。GrxS12 催化半胱氨酸的 pK(a) 非常低（3.9），使 GrxS12 本身容易受到 H(2)O(2)的氧化和亚硝基谷胱甘肽的直接谷胱甘肽化。谷胱甘肽化-GrxS12（GrxS12-SSG）暂时失去活性，直到被 GSH 去谷胱甘肽化。GrxS12 和谷胱甘肽之间的平衡（E(m(GrxS12-SSG))=-315 mV，pH 7.0）的特征是在 pH 7.0 时，K(ox) 值为 310，如在黑暗中的叶绿体中，以及在 pH 7.9 时，如在光照下的叶绿体中，K(ox) 值为 69。
根据热力学数据，GrxS12-SSG 预计会在 GSH 池轻度氧化的情况下在体内积累，这种情况可能在胁迫下发生。此外，GrxS12-SSG 在黑暗中的叶绿体中比在光照下更稳定。
这些特殊的催化和热力学性质可以使 GrxS12 作为一种与应激相关的氧化还原传感器发挥作用，从而使谷胱甘肽通过 GrxS12 靶蛋白的谷胱甘肽化发挥信号作用。
