Departments of Microbiology, Immunology and Molecular Genetics, University of California, Los Angeles, CA 90095-1489, USA.
Electrophoresis. 2009 Oct;30(20):3598-606. doi: 10.1002/elps.200900259.
A recently developed proteomic strategy, the "GG-azide"-labeling approach, is described for the detection and proteomic analysis of geranylgeranylated proteins. This approach involves metabolic incorporation of a synthetic azido-geranylgeranyl analog and chemoselective derivatization of azido-geranylgeranyl-modified proteins by the "click" chemistry, using a tetramethylrhodamine-alkyne. The resulting conjugated proteins can be separated by 1-D or 2-D and pH fractionation, and detected by fluorescence imaging. This method is compatible with downstream LC-MS/MS analysis. Proteomic analysis of conjugated proteins by this approach identified several known geranylgeranylated proteins as well as Rap2c, a novel member of the Ras family. Furthermore, prenylation of progerin in mouse embryonic fibroblast cells was examined using this approach, demonstrating that this strategy can be used to study prenylation of specific proteins. The "GG-azide"-labeling approach provides a new tool for the detection and proteomic analysis of geranylgeranylated proteins, and it can readily be extended to other post-translational modifications.
一种新开发的蛋白质组学策略，即“GG-叠氮化物”标记法，用于检测和蛋白质组学分析香叶基香叶基化蛋白。该方法涉及合成叠氮香叶基香叶基类似物的代谢掺入，以及使用四甲基罗丹明-炔通过“点击”化学对叠氮香叶基香叶基化修饰的蛋白质进行化学选择性衍生化。由此产生的缀合蛋白可以通过 1-D 或 2-D 和 pH 分级分离，并通过荧光成像检测。该方法与下游 LC-MS/MS 分析兼容。通过这种方法对缀合蛋白进行蛋白质组学分析鉴定了几种已知的香叶基香叶基化蛋白，以及 Ras 家族的新成员 Rap2c。此外，还使用该方法检查了小鼠胚胎成纤维细胞中 progerin 的 prenylation，表明该策略可用于研究特定蛋白质的 prenylation。“GG-叠氮化物”标记法为香叶基香叶基化蛋白的检测和蛋白质组学分析提供了一种新工具，并且可以很容易地扩展到其他翻译后修饰。
