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TMTpro-高效蛋白质组范围测定试剂
作者:小柯机器人 发布时间:2020/3/23 21:44:52

美国哈佛医学院Joao A. PauloSteven P. Gygi团队合作取得一项新成果。他们开发了异丁基-脯氨酸铵离子报告结构(TMTpro)试剂,这是一组同量异位标记质量标签,可同时对16个样品进行蛋白质组范围的测定。2020316日《自然-方法学》杂志发表了这一成果。

在一个蛋白质组规模的示例数据集中,他们比较了使用和不使用Torin1处理的三种常见细胞系(三个重复),每个重复定量了8800多种蛋白质(每个蛋白质含7.5个肽平均值),每个蛋白质组的分析时间仅为1.1h。最后,他们改善了热稳定性测定法,以检查用DMSO120μM星形孢菌素处理后的蛋白质组范围的熔解改变(5次重复后)。该试验仅在一个18小时的实验中就确定了星形孢菌素与228种细胞激酶的结合剂量分层。TMTpro试剂可进行复杂的实验设计-所有这16种样品基本上没有缺失值,而且定量完整性也没有损失。

据介绍,等压标记可同时对多个样品进行全蛋白质组表达测量。在此介绍了一组基于TMTpro的扩展的16种同量异位试剂。这些试剂具有与现有串联质谱标签试剂相似的特性,但具有更高的裂解效率和信号强度。

附:英文原文

Title: TMTpro reagents: a set of isobaric labeling mass tags enables simultaneous proteome-wide measurements across 16 samples

Author: Jiaming Li, Jonathan G. Van Vranken, Laura Pontano Vaites, Devin K. Schweppe, Edward L. Huttlin, Chris Etienne, Premchendar Nandhikonda, Rosa Viner, Aaron M. Robitaille, Andrew H. Thompson, Karsten Kuhn, Ian Pike, Ryan D. Bomgarden, John C. Rogers, Steven P. Gygi, Joao A. Paulo

Issue&Volume: 2020-03-16

Abstract: Isobaric labeling empowers proteome-wide expression measurements simultaneously across multiple samples. Here an expanded set of 16 isobaric reagents based on an isobutyl-proline immonium ion reporter structure (TMTpro) is presented. These reagents have similar characteristics to existing tandem mass tag reagents but with increased fragmentation efficiency and signal. In a proteome-scale example dataset, we compared eight common cell lines with and without Torin1 treatment with three replicates, quantifying more than 8,800 proteins (mean of 7.5 peptides per protein) per replicate with an analysis time of only 1.1h per proteome. Finally, we modified the thermal stability assay to examine proteome-wide melting shifts after treatment with DMSO, 1 or 20μM staurosporine with five replicates. This assay identified and dose-stratified staurosporine binding to 228 cellular kinases in just one, 18-h experiment. TMTpro reagents allow complex experimental designs—all with essentially no missing values across the 16 samples and no loss in quantitative integrity.

DOI: 10.1038/s41592-020-0781-4

Source: https://www.nature.com/articles/s41592-020-0781-4

期刊信息

Nature Methods:《自然—方法学》,创刊于2004年。隶属于施普林格·自然出版集团,最新IF:28.467
官方网址:https://www.nature.com/nmeth/
投稿链接:https://mts-nmeth.nature.com/cgi-bin/main.plex