Exponentially modified protein abundance index (emPAI) for estimation of absolute protein amount in proteomics by the number of sequenced peptides per protein
Exponentially modified protein abundance index (emPAI) for estimation of absolute protein amount in proteomics by the number of sequenced peptides per protein
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DOI：
10.1074/mcp.m500061-mcp200
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发表时间：
2005-09-01
影响因子：
7
通讯作者：
Mann, M
中科院分区：
文献类型：
--
作者：
Ishihama, Y;Oda, Y;Mann, M
为了估计复杂混合物中的绝对蛋白质含量，我们先前将蛋白质丰度指数（PAI）定义为观察到的肽的数量除以每个蛋白质可观察的肽数量（Rappsilber，J.，Ryder，U.，U.，Lamond，A。I.和A. I.和和Mann，M。（2002）人类剪接的大规模蛋白质组学分析。 1245）。在这里，我们报告说，在不同浓度的血清白蛋白下获得的PAI值与LC-MS/MS实验中蛋白质浓度的对数具有线性关系。小鼠全细胞裂解物中的46种蛋白也是这种情况。对于绝对定量，将PAI转换为指数修饰的PAI（EMPAI），等于10（pai）减去一个，该pai与蛋白质混合物中的蛋白质含量成正比。对于整个裂解物中的46种蛋白质，基于EMPAI的丰度与实际值的偏差百分比平均在63％以内，相似或更好，而不是通过蛋白质染色确定丰度。将EMPAI应用于人类癌细胞系中的基因和蛋白质表达之间的综合蛋白质表达分析，HCT116。 EMPAI的值很容易计算，并将重要的定量信息添加到蛋白质组学实验中；因此，我们建议应在大规模蛋白质组学识别项目中进行报告。
To estimate absolute protein contents in complex mixtures, we previously defined a protein abundance index (PAI) as the number of observed peptides divided by the number of observable peptides per protein (Rappsilber, J., Ryder, U., Lamond, A. I., and Mann, M. ( 2002) Large-scale proteomic analysis of the human spliceosome. Genome. Res. 12, 1231 - 1245). Here we report that PAI values obtained at different concentrations of serum albumin show a linear relationship with the logarithm of protein concentration in LC-MS/MS experiments. This was also the case for 46 proteins in a mouse whole cell lysate. For absolute quantitation, PAI was converted to exponentially modified PAI (emPAI), equal to 10(PAI) minus one, which is proportional to protein content in a protein mixture. For the 46 proteins in the whole lysate, the deviation percentages of the emPAI-based abundances from the actual values were within 63% on average, similar or better than determination of abundance by protein staining. emPAI was applied to comprehensive protein expression analysis and to a comparison study between gene and protein expression in a human cancer cell line, HCT116. The values of emPAI are easily calculated and add important quantitation information to proteomic experiments; therefore we suggest that they should be reported in large scale proteomic identification projects.