Quantification of Detergents Complexed with Membrane Proteins.
Quantification of Detergents Complexed with Membrane Proteins.
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DOI：
10.1038/srep41751
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发表时间：
2017-02-08
影响因子：
4.6
通讯作者：
Falson P
中科院分区：
文献类型：
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作者：
Chaptal V;Delolme F;Kilburg A;Magnard S;Montigny C;Picard M;Prier C;Monticelli L;Bornert O;Agez M;Ravaud S;Orelle C;Wagner R;Jawhari A;Broutin I;Pebay-Peyroula E;Jault JM;Kaback HR;le Maire M;Falson P
大多数膜蛋白的研究需要使用去污剂，但由于缺乏一个通用的，准确的和快速的方法来量化它们，许多不确定性仍然存在，阻碍了正确的功能和结构数据分析。为了解决这个问题，我们提出了一种基于基质辅助激光解吸/电离质谱（MALDI-TOF MS）的方法，该方法允许定量与膜蛋白复合的纯洗涤剂或混合洗涤剂。我们用各种洗涤剂和膜蛋白验证了该方法。我们将该过程自动化，从而允许广泛使用的常规定量。作为第一个例子，我们展示了如何获得与膜蛋白复合的洗涤剂的量的信息，这对于脂质体或纳米盘重构是必不可少的。由于该方法，我们还展示了如何可靠和容易地估计洗涤剂冠直径，并选择最小的尺寸，这对于有利于蛋白质-蛋白质接触和触发/促进膜蛋白质结晶至关重要，并可视化用于Cryo-EM研究的洗涤剂带。
Most membrane proteins studies require the use of detergents, but because of the lack of a general, accurate and rapid method to quantify them, many uncertainties remain that hamper proper functional and structural data analyses. To solve this problem, we propose a method based on matrix-assisted laser desorption/ionization mass spectrometry (MALDI-TOF MS) that allows quantification of pure or mixed detergents in complex with membrane proteins. We validated the method with a wide variety of detergents and membrane proteins. We automated the process, thereby allowing routine quantification for a broad spectrum of usage. As a first illustration, we show how to obtain information of the amount of detergent in complex with a membrane protein, essential for liposome or nanodiscs reconstitutions. Thanks to the method, we also show how to reliably and easily estimate the detergent corona diameter and select the smallest size, critical for favoring protein-protein contacts and triggering/promoting membrane protein crystallization, and to visualize the detergent belt for Cryo-EM studies.