A survey of the Arabidopsis thaliana mitochondrial phosphoproteome
A survey of the Arabidopsis thaliana mitochondrial phosphoproteome
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DOI：
10.1002/pmic.200900064
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发表时间：
2009-09-01
期刊：
影响因子：
3.4
通讯作者：
Heazlewood, Joshua L.
中科院分区：
文献类型：
--
作者：
Ito, Jun;Taylor, Nicolas L.;Heazlewood, Joshua L.
植物线粒体在细胞能量产生、新陈代谢和逆境反应中起着核心作用。最近对哺乳动物和酵母线粒体的磷酸蛋白质组学研究表明，蛋白质磷酸化可能是一种广泛的重要线粒体过程的调节机制。本研究利用亲和富集法和蛋白质组学方法研究了模式植物拟南芥细胞悬浮液中纯化的线粒体中的蛋白质磷酸化。用Pro-Q Diamond磷酸化凝胶内染料对拟南芥线粒体蛋白和亲和力增强的磷蛋白进行2-DE分离，检测到18种可能的磷酸化蛋白，包括线粒体代谢酶、热休克蛋白、一种蛋白酶和几种未知功能的蛋白。与酵母和小鼠的线粒体磷蛋白组的比较表明，这三个物种几乎没有经过验证的磷蛋白。用二氧化钛富集法和MS/MS对18种线粒体蛋白中的7种蛋白的磷酸化位点进行了表征。在此过程中，还发现了71个拟南芥蛋白的磷酸化多肽，它们不存在于线粒体中，但在不同类型的线粒体制剂中发现了污染物，表明与拟南芥的其他细胞成分相比，拟南芥线粒体成分的磷酸化水平较低。从这项研究中获得的信息为更好地理解拟南芥中亚细胞和细胞水平的蛋白质磷酸化提供了信息。
Plant mitochondria play central roles in cellular energy production, metabolism and stress responses. Recent phosphoproteomic studies in mammalian and yeast mitochondria have presented evidence indicating that protein phosphorylation is a likely regulatory mechanism across a broad range of important mitochondrial processes. This study investigated protein phosphorylation in purified mitochondria from cell suspensions of the model plant Arabidopsis thaliana using affinity enrichment and proteomic tools. Eighteen putative phospho-proteins consisting of mitochondrial metabolic enzymes, HSPs, a protease and several proteins of unknown function were detected on 2-DE separations of Arabidopsis mitochondrial proteins and affinity-enriched phosphoproteins using the Pro-Q Diamond phospho-specific in-gel dye. Comparisons with mitochondrial phosphoproteomes of yeast and mouse indicate that these three species share few validated phosphoproteins. Phosphorylation sites for seven of the eighteen mitochondrial proteins were characterized by titanium dioxide enrichment and MS/MS. In the process, 71 phosphopeptides from Arabidopsis proteins which are not present in mitochondria but found as contaminants in various types of mitochondrial preparations were also identified, indicating the low level of phosphorylation of mitochondrial components compared with other cellular components in Arabidopsis. Information gained from this study provides a better understanding of protein phosphorylation at both the subcellular and the cellular level in Arabidopsis.