Global Membrane Protein Interactome Analysis using In vivo Crosslinking and Mass Spectrometry-based Protein Correlation Profiling.
Global Membrane Protein Interactome Analysis using In vivo Crosslinking and Mass Spectrometry-based Protein Correlation Profiling.
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DOI：
10.1074/mcp.o115.055467
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发表时间：
2016-07
期刊：
影响因子：
--
通讯作者：
Lamond AI
中科院分区：
文献类型：
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作者：
Larance M;Kirkwood KJ;Tinti M;Brenes Murillo A;Ferguson MA;Lamond AI
我们提出了一种使用体内交联结合HPLC-MS的方法，通过蛋白质相关性分析对内源性蛋白质复合物进行全局分析。甲醛交联的蛋白质复合物提取高产量使用变性缓冲液，在色谱分离过程中保持复杂的溶解度。我们表明，这有效地检测完整的膜和膜相关蛋白复合物，除了可溶性复合物，允许识别和分析的复合物在本地提取物中不可访问。我们比较了在天然和甲醛交联的U2 OS细胞提取物中通过HPLC-MS蛋白质相关性分析检测到的蛋白质复合物。来自U2 OS细胞的体内交联和天然蛋白质复合物的这些蛋白质组范围的数据集可通过可搜索的在线数据库（www.peptracker.com/epd）免费获得。原始数据也可通过ProteomeXchange（标识符PXD 003754）获得。
We present a methodology using in vivo crosslinking combined with HPLC-MS for the global analysis of endogenous protein complexes by protein correlation profiling. Formaldehyde crosslinked protein complexes were extracted with high yield using denaturing buffers that maintained complex solubility during chromatographic separation. We show this efficiently detects both integral membrane and membrane-associated protein complexes,in addition to soluble complexes, allowing identification and analysis of complexes not accessible in native extracts. We compare the protein complexes detected by HPLC-MS protein correlation profiling in both native and formaldehyde crosslinked U2OS cell extracts. These proteome-wide data sets of both in vivo crosslinked and native protein complexes from U2OS cells are freely available via a searchable online database (www.peptracker.com/epd). Raw data are also available via ProteomeXchange (identifier PXD003754).