De novo protein crystal structure determination from X-ray free-electron laser data
De novo protein crystal structure determination from X-ray free-electron laser data
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DOI：
10.1038/nature12773
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发表时间：
2014-01-09
期刊：
影响因子：
64.8
通讯作者：
Schlichting, Ilme
中科院分区：
文献类型：
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作者：
Barends, Thomas R. M.;Foucar, Lutz;Schlichting, Ilme
蛋白质晶体结构的测定由于需要宏观晶体而受到阻碍。X射线自由电子激光器（FEL）提供飞秒持续时间的极强脉冲，这允许以“破坏前衍射”方法从纳米到微米尺寸的晶体（1-4）收集数据。到目前为止，使用FEL进行的所有蛋白质结构测定都是基于相关已知结构的先前知识（1-5）。在这里，我们表明，X射线自由电子激光数据可以用于从头蛋白质结构的测定，也就是说，没有以前的知识的结构。使用新兴的连续飞秒晶体学技术（1- 4，6），我们进行了单波长的异常散射测量的微晶体的成熟的模型系统溶菌酶，在复杂的镧系元素化合物。使用蒙特-卡罗积分（6，7），我们获得了高质量的衍射强度，从中可以确定实验相位，从而产生了足以自动构建蛋白质结构的实验电子密度图。这证明了使用FEL确定新蛋白质结构的可行性。我们预计，连续飞秒晶体学将成为一个重要的工具，用于蛋白质的结构确定是难以结晶，如膜蛋白（1，2，8）。
The determination of protein crystal structures is hampered by the need for macroscopic crystals. X-ray free-electron lasers (FELs) provide extremely intense pulses of femtosecond duration, which allow data collection from nanometre- to micrometre-sized crystals(1-4) in a 'diffraction-before-destruction' approach. So far, all protein structure determinations carried out using FELs have been based on previous knowledge of related, known structures(1-5). Here we show that X-ray FEL data can be used for de novo protein structure determination, that is, without previous knowledge about the structure. Using the emerging technique of serial femtosecond crystallography(1-4,6), we performed single-wavelength anomalous scattering measurements on microcrystals of the well-established model system lysozyme, in complex with a lanthanide compound. Using Monte-Carlo integration(6,7), we obtained high-quality diffraction intensities from which experimental phases could be determined, resulting in an experimental electron density map good enough for automated building of the protein structure. This demonstrates the feasibility of determining novel protein structures using FELs. We anticipate that serial femtosecond crystallography will become an important tool for the structure determination of proteins that are difficult to crystallize, such as membrane proteins(1,2,8).