Children's Hospital of Philadelphia Research Institute and Department of Pharmacology, Children's Hospital of Philadelphia and University of Pennsylvania, Philadelphia, PA 19104, USA.
Proc Natl Acad Sci U S A. 2010 Sep 28;107(39):16958-63. doi: 10.1073/pnas.1008036107. Epub 2010 Sep 13.
S-nitrosylation, the selective posttranslational modification of protein cysteine residues to form S-nitrosocysteine, is one of the molecular mechanisms by which nitric oxide influences diverse biological functions. In this study, unique MS-based proteomic approaches precisely pinpointed the site of S-nitrosylation in 328 peptides in 192 proteins endogenously modified in WT mouse liver. Structural analyses revealed that S-nitrosylated cysteine residues were equally distributed in hydrophobic and hydrophilic areas of proteins with an average predicted pK(a) of 10.01 ± 2.1. S-nitrosylation sites were over-represented in α-helices and under-represented in coils as compared with unmodified cysteine residues in the same proteins (χ(2) test, P < 0.02). A quantile-quantile probability plot indicated that the distribution of S-nitrosocysteine residues was skewed toward larger surface accessible areas compared with the unmodified cysteine residues in the same proteins. Seventy percent of the S-nitrosylated cysteine residues were surrounded by negatively or positively charged amino acids within a 6-Å distance. The location of cysteine residues in α-helices and coils in highly accessible surfaces bordered by charged amino acids implies site directed S-nitrosylation mediated by protein-protein or small molecule interactions. Moreover, 13 modified cysteine residues were coordinated with metals and 15 metalloproteins were endogenously modified supporting metal-catalyzed S-nitrosylation mechanisms. Collectively, the endogenous S-nitrosoproteome in the liver has structural features that accommodate multiple mechanisms for selective site-directed S-nitrosylation.
S-亚硝基化作用，即蛋白质半胱氨酸残基选择性的翻译后修饰形成 S-亚硝基化半胱氨酸，是一氧化氮影响多种生物功能的分子机制之一。在这项研究中，独特的基于 MS 的蛋白质组学方法精确地确定了 WT 小鼠肝脏中内源性修饰的 192 种蛋白质中的 328 种肽中的 S-亚硝基化位点。结构分析表明，S-亚硝基化半胱氨酸残基在蛋白质的疏水区和亲水区域均匀分布，平均预测 pK(a)为 10.01±2.1。与同一蛋白质中未修饰的半胱氨酸残基相比，S-亚硝基化位点在α-螺旋中过度表达，在卷曲中表达不足（卡方检验，P<0.02）。分位数-分位数概率图表明，与同一蛋白质中未修饰的半胱氨酸残基相比，S-亚硝基化半胱氨酸残基的分布偏向于更大的表面可及性区域。70%的 S-亚硝基化半胱氨酸残基在 6Å 距离内被带正电荷或负电荷的氨基酸所包围。高度可及表面边界处的α-螺旋和卷曲中的半胱氨酸残基的位置表明，由蛋白质-蛋白质或小分子相互作用介导的定点 S-亚硝基化。此外，有 13 个修饰的半胱氨酸残基与金属配位，15 种金属蛋白酶内源性修饰支持金属催化的 S-亚硝基化机制。总的来说，肝脏中的内源性 S-亚硝基化蛋白质组具有容纳多种选择性定点 S-亚硝基化机制的结构特征。
