Mechanism of Sirt1 NAD+-dependent Protein Deacetylase Inhibition by Cysteine S-Nitrosation
Mechanism of Sirt1 NAD+-dependent Protein Deacetylase Inhibition by Cysteine S-Nitrosation
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DOI：
10.1074/jbc.m116.754655
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发表时间：
2016-12-02
影响因子：
4.8
通讯作者：
Smith, Brian C.
中科院分区：
文献类型：
--
作者：
Kalous, Kelsey S.;Wynia-Smith, Sarah L.;Smith, Brian C.
沉默调节蛋白家族催化酰基赖氨酸残基的NAD（+）依赖性脱酰。人类编码七种sirtuins（Sirt 1 -7），最近的研究表明，半胱氨酸S-亚硝基化的Sirt 1的翻译后修饰与Sirt 1脱乙酰基酶底物的乙酰化增加相关。然而，S-亚硝化抑制Sirt 1的机制尚不清楚。在这里，我们表明，Sirt 1是transnitrosated和生理相关的亚硝基巯基S-亚硝基谷胱甘肽抑制。稳态动力学分析和结合测定与Sirt 1 S-亚硝化抑制NAD（+）和乙酰基赖氨酸底物的结合一致。Sirt 1的S-亚硝化与锌2+释放从保守的沉默调节蛋白锌2 +-四硫醇盐和损失的螺旋结构没有整体的热不稳定的酶。分子动力学模拟表明，由于Sirt 1 S-亚硝化导致的Zn 2+损失导致四硫醇盐亚结构域远离催化结构域的其余部分重新定位，从而破坏NAD（+）和乙酰基-赖氨酸结合位点。Sirt 1的S-亚硝化被逆转后，暴露于巯基为基础的还原剂，包括生理相关浓度的细胞还原剂谷胱甘肽。S-亚硝化的抑制导致Sirt 1活性的完全恢复，只有在Zn 2+的存在下，与作为Sirt 1抑制S-亚硝基谷胱甘肽治疗后的主要来源的Zn 2 +-四硫醇的S-亚硝化一致。
The sirtuin family of proteins catalyze the NAD(+)-dependent deacylation of acyl-lysine residues. Humans encode seven sirtuins (Sirt1-7), and recent studies have suggested that post-translational modification of Sirt1 by cysteine S-nitrosation correlates with increased acetylation of Sirt1 deacetylase substrates. However, the mechanism of Sirt1 inhibition by S-nitrosation was unknown. Here, we show that Sirt1 is transnitrosated and inhibited by the physiologically relevant nitrosothiol S-nitrosoglutathione. Steady-state kinetic analyses and binding assays were consistent with Sirt1 S-nitrosation inhibiting binding of both the NAD(+) and acetyl-lysine substrates. Sirt1 S-nitrosation correlated with Zn2+ release from the conserved sirtuin Zn2+-tetrathiolate and a loss of -helical structure without overall thermal destabilization of the enzyme. Molecular dynamics simulations suggested that Zn2+ loss due to Sirt1 S-nitrosation results in repositioning of the tetrathiolate subdomain away from the rest of the catalytic domain, thereby disrupting the NAD(+) and acetyl-lysine-binding sites. Sirt1 S-nitrosation was reversed upon exposure to the thiol-based reducing agents, including physiologically relevant concentrations of the cellular reducing agent glutathione. Reversal of S-nitrosation resulted in full restoration of Sirt1 activity only in the presence of Zn2+, consistent with S-nitrosation of the Zn2+-tetrathiolate as the primary source of Sirt1 inhibition upon S-nitrosoglutathione treatment.