Structure determination of an integral membrane protein at room temperature from crystals in situ.
Structure determination of an integral membrane protein at room temperature from crystals in situ.
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DOI：
10.1107/s139900471500423x
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发表时间：
2015-06
期刊：
影响因子：
--
通讯作者：
Alguel Y
中科院分区：
文献类型：
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作者：
Axford D;Foadi J;Hu NJ;Choudhury HG;Iwata S;Beis K;Evans G;Alguel Y
利用在室温下原位测量的同步辐射衍射数据，对完整的膜蛋白进行了X射线结构测定。本文介绍了直接在蒸气扩散结晶板上(原位)采集的同步辐射X射线衍射法测定完整膜蛋白结构的方法。就地暴露晶体可避免手动处理样品，而且由于是在室温下进行的，因此消除了冷冻保护的复杂性和样品冷冻引起的潜在结构异常。这种方法的关键是能够通过记录来自许多样本的每个样本的少量数据，然后使用专门的软件汇编产生的数据集来限制辐射损害。该方法的有效性是通过对流感嗜血杆菌Teha在2.3 á分辨率下的结构测定来确定的。该方法为利用第三代同步加速器光束线在室温下快速有效地确定膜蛋白质结构提供了一种有效的方法。
The X-ray structure determination of an integral membrane protein using synchrotron diffraction data measured in situ at room temperature is demonstrated. The structure determination of an integral membrane protein using synchrotron X-ray diffraction data collected at room temperature directly in vapour-diffusion crystallization plates (in situ) is demonstrated. Exposing the crystals in situ eliminates manual sample handling and, since it is performed at room temperature, removes the complication of cryoprotection and potential structural anomalies induced by sample cryocooling. Essential to the method is the ability to limit radiation damage by recording a small amount of data per sample from many samples and subsequently assembling the resulting data sets using specialized software. The validity of this procedure is established by the structure determination of Haemophilus influenza TehA at 2.3 Å resolution. The method presented offers an effective protocol for the fast and efficient determination of membrane-protein structures at room temperature using third-generation synchrotron beamlines.