Proteomic analysis of organ-specific post-translational lysine acetylation and -methylation in mice by use of anti-acetyllysine and -methyllysine mouse monoclonal antibodies
Proteomic analysis of organ-specific post-translational lysine acetylation and -methylation in mice by use of anti-acetyllysine and -methyllysine mouse monoclonal antibodies
复制标题
DOI：
10.1002/pmic.200500042
复制
发表时间：
2005-12-01
期刊：
影响因子：
3.4
通讯作者：
Komatsu, Y
中科院分区：
文献类型：
--
作者：
Iwabata, H;Yoshida, M;Komatsu, Y
翻译后赖氨酸乙酰化和甲基化是蛋白质中赖氨酸残基的两种主要翻译后修饰。最近，我们制备了泛反应性抗乙酰赖氨酸小鼠单克隆抗体，它能在各种氨基酸序列环境下结合N - ε -乙酰化赖氨酸残基。在本研究中，我们制备了与抗乙酰赖氨酸抗体相当的泛反应性抗甲基赖氨酸小鼠单克隆抗体。通过使用这些抗乙酰赖氨酸和抗甲基赖氨酸抗体，我们发现小鼠器官中赖氨酸乙酰化和甲基化蛋白质的模式在不同器官之间存在极大差异。我们选择大脑和骨骼肌作为模型案例，通过二维电泳（2 - DE）和随后的蛋白质印迹（Western blotting）进一步分析。在大脑中，发现α -微管蛋白在其基础水平上被高度乙酰化；并且α -烯醇化酶被证明是一种新发现的可能被乙酰化的蛋白质。神经丝轻链（NF - L）蛋白、热休克蛋白70（Hsc70）、α -微管蛋白片段、β -肌动蛋白和脑型肌酸激酶被鉴定为小鼠大脑中可能的赖氨酸甲基化蛋白质。在骨骼肌中，发现α -肌动蛋白的赖氨酸甲基化以及肌肉型肌酸激酶的赖氨酸乙酰化和甲基化是新的可能的赖氨酸修饰蛋白质。这里介绍的方法可能有助于发现仅使用传统蛋白质组学方法无法注意到的新型疾病标志物和/或药物靶分子。
Post-translational lysine-acetylation and -methylation are two major PTMs of lysine residues in proteins. Recently, we established pan-reactive anti-acetyllysine mouse mAbs, which can bind to N epsilon-acetylated lysine residues in various contexts of amino acid sequences. In the present study, we established pan-reactive anti-methyllysine mouse mAbs comparable to the anti-acetyllysine ones. By using these anti-acetyllysine and -methyllysine antibodies, we found that the pattern of lysine-acetylated and -methylated proteins in mouse organs showed extreme variation from organ to organ. We selected brain and skeletal muscle as model cases to be further analyzed by 2-DE followed by Westem blotting. In brain, alpha-tubulin at its basal level was found to be extremely acetylated; and alpha-enolase was shown to be a newly recognized possibly acetylated protein. NF-L protein, Hsc70, alpha-tubulin fragments, beta-actin, and brain-type creatine kinase were identified as putative lysine-methylated proteins in mouse brain. In skeletal muscle, lysine-methylation of alpha-actin and both lysine-acetylation and -methylation of muscle-type creatine kinase were found as novel putative lysine-modified proteins. The approach presented here might be useful to find novel disease markers and/or drug target molecules that would not be noticed by use of the traditional proteomic approach only.