Nass Karol, Bacellar Camila, Cirelli Claudio, Dworkowski Florian, Gevorkov Yaroslav, James Daniel, Johnson Philip J M, Kekilli Demet, Knopp Gregor, Martiel Isabelle, Ozerov Dmitry, Tolstikova Alexandra, Vera Laura, Weinert Tobias, Yefanov Oleksandr, Standfuss Jörg, Reiche Sven, Milne Christopher J
Paul Scherrer Institut, Forschungstrasse 111, Villigen 5232, Switzerland.
Center for Free-Electron Laser Science, Deutsches Elektronen-Synchrotron DESY, Notkestrasse 85, Hamburg 22607, Germany.
IUCrJ. 2021 Sep 23;8(Pt 6):905-920. doi: 10.1107/S2052252521008046. eCollection 2021 Nov 1.
Serial femtosecond crystallography (SFX) at X-ray free-electron lasers (XFELs) enables essentially radiation-damage-free macromolecular structure determination using microcrystals that are too small for synchrotron studies. However, SFX experiments often require large amounts of sample in order to collect highly redundant data where some of the many stochastic errors can be averaged out to determine accurate structure-factor amplitudes. In this work, the capability of the Swiss X-ray free-electron laser (SwissFEL) was used to generate large-bandwidth X-ray pulses [Δλ/λ = 2.2% full width at half-maximum (FWHM)], which were applied in SFX with the aim of improving the partiality of Bragg spots and thus decreasing sample consumption while maintaining the data quality. Sensitive data-quality indicators such as anomalous signal from native thaumatin micro-crystals and phasing results were used to quantify the benefits of using pink X-ray pulses to obtain accurate structure-factor amplitudes. Compared with data measured using the same setup but using X-ray pulses with typical quasi-monochromatic XFEL bandwidth (Δλ/λ = 0.17% FWHM), up to fourfold reduction in the number of indexed diffraction patterns required to obtain similar data quality was achieved. This novel approach, pink-beam SFX, facilitates the yet underutilized structure determination of challenging proteins at XFELs, thereby opening the door to more scientific breakthroughs.
利用X射线自由电子激光（XFEL）进行的连续飞秒晶体学（SFX），能够使用对于同步加速器研究而言太小的微晶来确定基本无辐射损伤的大分子结构。然而，SFX实验通常需要大量样品，以便收集高度冗余的数据，其中许多随机误差中的一些可以被平均掉，从而确定准确的结构因子振幅。在这项工作中，瑞士X射线自由电子激光（SwissFEL）的能力被用于产生大带宽X射线脉冲[Δλ/λ = 2.2%半高全宽（FWHM）]，这些脉冲被应用于SFX，目的是提高布拉格斑点的完整性，从而在保持数据质量的同时减少样品消耗。诸如天然奇异果甜蛋白微晶的反常信号和相位结果等敏感的数据质量指标，被用于量化使用粉红X射线脉冲获得准确结构因子振幅的益处。与使用相同装置但使用具有典型准单色XFEL带宽（Δλ/λ = 0.17% FWHM）的X射线脉冲测量的数据相比，在获得相似数据质量所需的索引衍射图案数量上实现了高达四倍的减少。这种新颖的方法，即粉红光束SFX，有助于在XFEL上对具有挑战性的蛋白质进行尚未充分利用的结构测定，从而为更多的科学突破打开了大门。
