The cardiac acetyl-lysine proteome.
The cardiac acetyl-lysine proteome.
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DOI：
10.1371/journal.pone.0067513
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发表时间：
2013
期刊：
影响因子：
3.7
通讯作者：
O'Rourke B
中科院分区：
文献类型：
--
作者：
Foster DB;Liu T;Rucker J;O'Meally RN;Devine LR;Cole RN;O'Rourke B
在心脏中，赖氨酸乙酰化与从病理重塑的转录控制到热量限制引起的心脏保护等过程有关。鉴于这种翻译后修饰的重要性日益凸显，我们采用蛋白质组学方法，利用豚鼠模型研究了赖氨酸乙酰化在心脏中的更广泛作用。简而言之，在蛋白水解和乙酰化肽的亲和力富集之前，将心脏分级为富含肌丝、线粒体和胞质溶胶的级分。 LC-MS/MS 分析鉴定出 1075 个乙酰化肽，包含 994 个乙酰化位点，映射到 240 个蛋白质，总体蛋白质错误发现率 <0.8%。线粒体靶标占已识别蛋白质的 59% 和位点的 64%。大多数乙酰蛋白是参与脂肪酸代谢、氧化磷酸化或 TCA 循环的酶。在胞质部分中，糖酵解、脂肪酸合成和脂质结合酶是突出的。核靶标包括组蛋白和转录调节因子 E1A(p300) 和 CREB 结合蛋白。我们的数据集与之前三项全球乙酰组学研究的比较独特地揭示了 53 种赖氨酸乙酰化蛋白质。具体来说，新鉴定的乙酰蛋白包括 Ca2+ 处理蛋白、RyR2 和 SERCA2，以及肌丝蛋白、肌球蛋白重链、肌球蛋白轻链和肌钙蛋白复合物亚基等。这些观察结果通过抗乙酰赖氨酸免疫印迹得到证实。总之，心脏赖氨酸乙酰化可能在心脏底物选择、生物能性能和氧化还原平衡维持中发挥作用。新的站点提出了许多潜在的机制，通过这些机制也可以调节兴奋-收缩耦合。
In the heart, lysine acetylation has been implicated in processes ranging from transcriptional control of pathological remodeling, to cardioprotection arising from caloric restriction. Given the emerging importance of this post-translational modification, we used a proteomic approach to investigate the broader role of lysine acetylation in the heart using a guinea pig model. Briefly, hearts were fractionated into myofilament-, mitochondrial- and cytosol-enriched fractions prior to proteolysis and affinity-enrichment of acetylated peptides. LC-MS/MS analysis identified 1075 acetylated peptides, harboring 994 acetylation sites that map to 240 proteins with a global protein false discovery rate <0.8%. Mitochondrial targets account for 59% of identified proteins and 64% of sites. The majority of the acetyl-proteins are enzymes involved in fatty acid metabolism, oxidative phosphorylation or the TCA cycle. Within the cytosolic fraction, the enzymes of glycolysis, fatty acid synthesis and lipid binding are prominent. Nuclear targets included histones and the transcriptional regulators E1A(p300) and CREB binding protein. Comparison of our dataset with three previous global acetylomic studies uniquely revealed 53 lysine-acetylated proteins. Specifically, newly-identified acetyl-proteins include Ca2+-handling proteins, RyR2 and SERCA2, and the myofilament proteins, myosin heavy chain, myosin light chains and subunits of the Troponin complex, among others. These observations were confirmed by anti-acetyl-lysine immunoblotting. In summary, cardiac lysine acetylation may play a role in cardiac substrate selection, bioenergetic performance, and maintenance of redox balance. New sites suggest a host of potential mechanisms by which excitation-contraction coupling may also be modulated.
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