Temperature-dependent radiation sensitivity and order of 70S ribosome crystals
Temperature-dependent radiation sensitivity and order of 70S ribosome crystals
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温度依赖性辐射敏感性和 70S 核糖体晶体的顺序
DOI：
10.1107/s1399004714017672
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发表时间：
2014
期刊：
影响因子：
--
通讯作者：
Thorne, Robert E.
中科院分区：
文献类型：
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作者：
Warkentin, Matthew;Hopkins, Jesse B.;Haber, Jonah B.;Blaha, Gregor;Thorne, Robert E.
迄今为止的所有证据表明，在T = 100 K时，所有蛋白质晶体表现出相当的敏感性，X射线损伤时，使用全球度量，如缩放B因子或积分强度与剂量的变化进行量化。 这与冷冻电子显微镜的观察结果一致，因为几乎所有的蛋白质和溶剂的扩散运动，包括辐射损伤引起的运动，都被冻结了。但是，在室温下，辐射诱导的自由基可以自由扩散，蛋白质和晶格结构可以自由松弛以应对局部损伤，不同蛋白质的敏感性如何比较？可以预期的是，具有广泛构象自由度的大复合物将比小的紧凑的球状蛋白质对辐射更敏感。作为一个测试的情况下，70 S核糖体晶体的辐射敏感性已被检查。在T = 100和300 K时，半剂量分别为64 MGy（3 μ m分辨率）和150 kGy（5 μ m分辨率）。 晶体学实验中的最大耐受剂量取决于初始分辨率或所需分辨率。当考虑到初始数据集分辨率的差异时，前半剂量与模型蛋白质的半剂量大致一致，100/300 K的半剂量比大约大10倍。 70 S核糖体晶体在100 K下的分辨率相对于300 K显著增加，这是由于冷却诱导的有序化，而不是由于辐射敏感性降低和辐射损伤较慢。
All evidence to date indicates that at T = 100 K all protein crystals exhibit comparable sensitivity to X-ray damage when quantified using global metrics such as change in scaling B factor or integrated intensity versus dose. This is consistent with observations in cryo-electron microscopy, and results because nearly all diffusive motions of protein and solvent, including motions induced by radiation damage, are frozen out. But how do the sensitivities of different proteins compare at room temperature, where radiation-induced radicals are free to diffuse and protein and lattice structures are free to relax in response to local damage? It might be expected that a large complex with extensive conformational degrees of freedom would be more radiation sensitive than a small, compact globular protein. As a test case, the radiation sensitivity of 70S ribosome crystals has been examined. At T = 100 and 300 K, the half doses are 64 MGy (at 3 Å resolution) and 150 kGy (at 5 Å resolution), respectively. The maximum tolerable dose in a crystallography experiment depends upon the initial or desired resolution. When differences in initial data-set resolution are accounted for, the former half dose is roughly consistent with that for model proteins, and the 100/300 K half-dose ratio is roughly a factor of ten larger. 70S ribosome crystals exhibit substantially increased resolution at 100 K relative to 300 K owing to cooling-induced ordering and not to reduced radiation sensitivity and slower radiation damage.