Phosphoproteomic analysis of the mouse brain cytosol reveals a predominance of protein phosphorylation in regions of intrinsic sequence disorder
Phosphoproteomic analysis of the mouse brain cytosol reveals a predominance of protein phosphorylation in regions of intrinsic sequence disorder
复制标题
DOI：
10.1074/mcp.m700564-mcp200
复制
发表时间：
2008-07-01
影响因子：
7
通讯作者：
Choudhary, Jyoti S.
中科院分区：
文献类型：
--
作者：
Collins, Mark O.;Yu, Lu;Choudhary, Jyoti S.
我们分析了小鼠前脑胞浆磷酸化蛋白质组使用顺序（蛋白质和肽）IMAC纯化，酶去磷酸化，有针对性的串联质谱分析策略。总的来说，使用互补的磷酸富集和LCMS/MS策略，512磷酸化位点的540个非冗余磷酸肽从162胞质磷酸蛋白的特点。蛋白质结构域和氨基酸序列组成的分析，这一数据集的胞质磷蛋白显示，它是显着丰富的内在序列紊乱，这种富集与细胞位置和磷酸化状态。MS发现的大多数磷酸化位点位于结构蛋白结构域之外（97%），但大多数位于内在序列紊乱区域（86%）。368个磷酸化位点位于长的紊乱区域（超过40个氨基酸长），并且94%的蛋白质含有至少一个这样的长的紊乱区域。此外，我们发现在这个数据集中的58个磷酸化位点发生在14-3-3结合共有基序，线性基序与蛋白质中的非结构化区域相关。这些结果表明，在这个数据集中，蛋白磷酸化显着耗尽的蛋白质结构域和显着丰富的无序蛋白质序列和丰富的内在序列紊乱可能是磷酸化蛋白质组的一个共同特征。这支持了蛋白质中的无序区域允许激酶、磷酸酶和磷酸化依赖性结合蛋白获得靶序列以调节局部蛋白质构象和活性的假设。
We analyzed the mouse forebrain cytosolic phosphoproteome using sequential (protein and peptide) IMAC purifications, enzymatic dephosphorylation, and targeted tandem mass spectrometry analysis strategies. In total, using complementary phosphoenrichment and LCMS/MS strategies, 512 phosphorylation sites on 540 non-redundant phosphopeptides from 162 cytosolic phospho-proteins were characterized. Analysis of protein domains and amino acid sequence composition of this data set of cytosolic phosphoproteins revealed that it is significantly enriched in intrinsic sequence disorder, and this enrichment is associated with both cellular location and phosphorylation status. The majority of phosphorylation sites found by MS were located outside of structural protein domains (97%) but were mostly located in regions of intrinsic sequence disorder (86%). 368 phosphorylation sites were located in long regions of disorder (over 40 amino acids long), and 94% of proteins contained at least one such long region of disorder. In addition, we found that 58 phosphorylation sites in this data set occur in 14-3-3 binding consensus motifs, linear motifs that are associated with unstructured regions in proteins. These results demonstrate that in this data set protein phosphorylation is significantly depleted in protein domains and significantly enriched in disordered protein sequences and that enrichment of intrinsic sequence disorder may be a common feature of phosphoproteomes. This supports the hypothesis that disordered regions in proteins allow kinases, phosphatases, and phosphorylation-dependent binding proteins to gain access to target sequences to regulate local protein conformation and activity.