Drop-on-demand sample delivery for studying biocatalysts in action at X-ray free-electron lasers.
Drop-on-demand sample delivery for studying biocatalysts in action at X-ray free-electron lasers.
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DOI：
10.1038/nmeth.4195
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发表时间：
2017-04
期刊：
影响因子：
48
通讯作者：
Yano J
中科院分区：
文献类型：
--
作者：
Fuller FD;Gul S;Chatterjee R;Burgie ES;Young ID;Lebrette H;Srinivas V;Brewster AS;Michels-Clark T;Clinger JA;Andi B;Ibrahim M;Pastor E;de Lichtenberg C;Hussein R;Pollock CJ;Zhang M;Stan CA;Kroll T;Fransson T;Weninger C;Kubin M;Aller P;Lassalle L;Bräuer P;Miller MD;Amin M;Koroidov S;Roessler CG;Allaire M;Sierra RG;Docker PT;Glownia JM;Nelson S;Koglin JE;Zhu D;Chollet M;Song S;Lemke H;Liang M;Sokaras D;Alonso-Mori R;Zouni A;Messinger J;Bergmann U;Boal AK;Bollinger JM Jr;Krebs C;Högbom M;Phillips GN Jr;Vierstra RD;Sauter NK;Orville AM;Kern J;Yachandra VK;Yano J
X射线自由电子激光(XFEL)源的X射线结晶学是在生物相关温度下研究大分子的有力手段。此外，当与X射线发射光谱(XES)等补充技术相结合时，可以同时获得金属酶的全局结构和化学性质，为了解蛋白质结构/动力学和活性部位的化学之间的相互作用提供了新的见解。实现这样的多模式方法可能会受到优化每个单独方法的冲突要求的影响。特别是，样本传递的方法对数据质量有很大影响。我们在这里提出了一种新的、坚固的方法，使用声学液滴喷射结合传送带驱动来提供受控样本量，传送带驱动针对广泛时间范围内的光化学和化学反应的结晶学和光谱测量进行了优化。对光系统II、光敏色素感受器和核糖核苷酸还原酶R2的研究表明了这种方法的强大和多功能性。
X-ray crystallography at X-ray free-electron laser (XFEL) sources is a powerful method for studying macromolecules at biologically relevant temperatures. Moreover, when combined with complementary techniques like X-ray emission spectroscopy (XES), both global structures and chemical properties of metalloenzymes can be obtained concurrently, providing new insights into the interplay between the protein structure/dynamics and chemistry at an active site. Implementing such a multimodal approach can be compromised by conflicting requirements to optimize each individual method. In particular, the method used for sample delivery greatly impacts the data quality. We present here a new, robust way of delivering controlled sample amounts on demand using acoustic droplet ejection coupled with a conveyor belt drive that is optimized for crystallography and spectroscopy measurements of photochemical and chemical reactions over a wide range of time scales. Studies with photosystem II, the phytochrome photoreceptor, and ribonucleotide reductase R2 illustrate the power and versatility of this method.