Formation of a Stabilized Cysteine Sulfinic Acid Is Critical for the Mitochondrial Function of the Parkinsonism Protein DJ-1
Formation of a Stabilized Cysteine Sulfinic Acid Is Critical for the Mitochondrial Function of the Parkinsonism Protein DJ-1
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DOI：
10.1074/jbc.m806599200
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发表时间：
2009-03-06
影响因子：
4.8
通讯作者：
Wilson, Mark A.
中科院分区：
文献类型：
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作者：
Blackinton, Jeff;Lakshminarasimhan, Mahadevan;Wilson, Mark A.
半胱氨酸-亚磺酸的形成最近被认为是一种将蛋白质功能与细胞氧化状态联系起来的修饰。人类DJ-1是一种与遗传性帕金森病相关的蛋白质，它很容易在一个保守的半胱氨酸残基（人类DJ-1中的Cys(106)）上形成半胱氨酸亚磺酸。Cys（106）的突变导致该蛋白在细胞培养和模式生物中失去其正常的保护功能。然而，尚不清楚这些突变体中DJ-1保护功能的丧失是由于缺少Cys（106）氧化还是由于缺少半胱氨酸残基本身。为了解决这个问题，我们在人类DJ-1的近端谷氨酸残基（Glu(18)）上设计了一系列取代，通过改变氢键来改变Cys（106）的氧化倾向。我们发现两个突变，E18N和E18Q，允许Cys（106）在温和的条件下被氧化成Cys(106)-亚磺酸。相反，E18D突变稳定了半胱氨酸-亚磺酸，在溶液和体内都很容易还原为硫醇。通过线粒体分裂和细胞活力测定，我们发现E18N和E18Q都可以部分替代野生型DJ-1。相反，氧化受损的E18D突变体表现为无活性的C106A突变体，不能保护细胞。因此，我们得出结论，Cys(106)-亚磺酸的形成是调节DJ-1保护功能的关键修饰。
The formation of cysteine-sulfinic acid has recently become appreciated as a modification that links protein function to cellular oxidative status. Human DJ-1, a protein associated with inherited parkinsonism, readily forms cysteine-sulfinic acid at a conserved cysteine residue (Cys(106) in human DJ-1). Mutation of Cys(106) causes the protein to lose its normal protective function in cell culture and model organisms. However, it is unknown whether the loss of DJ-1 protective function in these mutants is due to the absence of Cys(106) oxidation or the absence of the cysteine residue itself. To address this question, we designed a series of substitutions at a proximal glutamic acid residue (Glu(18)) in human DJ-1 that alter the oxidative propensity of Cys(106) through changes in hydrogen bonding. We show that two mutations, E18N and E18Q, allow Cys(106) to be oxidized to Cys(106)-sulfinic acid under mild conditions. In contrast, the E18D mutation stabilizes a cysteine-sulfenic acid that is readily reduced to the thiol in solution and in vivo. We show that E18N and E18Q can both partially substitute for wild-type DJ-1 using mitochondrial fission and cell viability assays. In contrast, the oxidatively impaired E18D mutant behaves as an inactive C106A mutant and fails to protect cells. We therefore conclude that formation of Cys(106)-sulfinic acid is a key modification that regulates the protective function of DJ-1.