Toward chromatographic analysis of interacting protein networks
Toward chromatographic analysis of interacting protein networks
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DOI：
10.1016/j.chroma.2007.10.067
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发表时间：
2008-01-12
影响因子：
4.1
通讯作者：
Regnier, Fred
中科院分区：
文献类型：
--
作者：
Liu, Xiuping;Yang, Wen-chu;Regnier, Fred
蛋白质复合物，统称为细胞相互作用组，似乎在细胞调节中发挥重要作用。目前认为，相互作用体可能由数百个蛋白质组装体组成。本文所述工作的目的是研究广泛用于制备性分离天然蛋白质的色谱方法可以以这样一种方式并入全球蛋白质组学方法的前景，即具有足够稳定性以经受色谱的蛋白质复合物的一级结构可以沿着它们参与蛋白质复合物。因为大小的广泛差异是蛋白质复合物的独特特征，所以将尺寸排阻色谱法（SEC）并入所检查的所有分级分离策略中。阴离子交换层析（AEC）和疏水相互作用层析（HIC）也进行了检查，因为这些方法在制备具有天然结构的蛋白质中显示出广泛的实用性。在酵母裂解物中鉴定的所有蛋白质中，略多于三分之一的蛋白质被发现从SEC、AEC和fHC柱中分离，其表观分子量远高于从其亲本基因中预测的分子量。这些结果被解释为意味着这些蛋白质作为蛋白质复合物的组分通过柱迁移。基于使用多维SEC -> RPLC（反相液相色谱）、AEC -> SEC和HIC -> SEC系统的研究，可以得出结论，当至少一个分级维度包括天然蛋白质的SEC时，复合物中蛋白质的识别可以容易地并入用于全局蛋白质组学的多维色谱方法中。(c)2007 Elsevier B. V.保留所有权利。
Protein complexes, collectively referred to as the cellular interactome, appear to play a major role in cellular regulation. At present it is thought that the interactome could be composed of hundreds of protein assemblies. The objective of the work described here was to examine the prospect that chromatographic methods widely used in the preparative isolation of native proteins could be incorporated into global proteomics methods in such a way that the primary structure of protein complexes of sufficient stability to survive chromatography could be recognized along with their participation in protein complexes. Because wide differences in sizes are a unique feature of protein complexes, size-exclusion chromatography (SEC) was incorporated into all the fractionation strategies examined. Anion-exchange chromatography (AEC) and hydrophobic-interaction chromatography (HIC) were also examined because of the broad utility that these methods have shown in the preparation of proteins with native structure. Slightly more than a third of all proteins identified in yeast lysates were found to elute from SEC, AEC, and fHC columns with an apparent molecular weight much higher than that predicted from their parent gene. These results were interpreted to mean that these proteins were migrating through columns as components of protein complexes. Based on studies with multidimensional SEC -> RPLC (reversed-phase liquid chromatography), AEC -> SEC, and HIC -> SEC systems, it was concluded that recognition of proteins in complexes could be easily incorporated into multidimensional chromatographic methods for global proteomics when at least one of the fractionation dimensions included SEC of native proteins. (c) 2007 Elsevier B.V. All rights reserved.