Khakurel Krishna Prasad, Hosomi Kei, Inami Wataru, Yoshimasa Kawata
Extreme Light Infrastructure (ERIC), Za Radnici 835, 25241Dolni Brezany, Czechia.
Research Institute of Electronics Shizuoka University 3-5-1 Johoku, Chuo-ku Hamamatsu432-8561 Japan.
J Appl Crystallogr. 2024 Nov 22;57(Pt 6):1907-1912. doi: 10.1107/S1600576724010124. eCollection 2024 Dec 1.
Crystallography has been the routine technique for studying high-resolution structures of proteins for over five decades. A major bottleneck in structure determination of macromolecules is obtaining crystals of a size and quality suitable for single-crystal X-ray crystallography experiments. Many challenging proteins either fail to grow into crystals or fail to grow into crystals of a size suitable for obtaining high-resolution structures using conventional X-ray crystallography. When it comes to smaller crystals, they can be used either for seeding to get larger crystals or for serial crystallography and electron diffraction for obtaining the structures. For both purposes, a limiting step is to non-invasively image such small crystals of sub-micrometre dimensions and to screen the conditions where such crystals prevail. Here we use cathodoluminescence-based (CL-based) nanoscopy to image protein nanocrystals. We show that crystals of micrometre and submicrometre dimensions can be non-invasively imaged by the CL-based nanoscope. The results presented here demonstrate the feasibility of non-invasive imaging of protein crystals with sub-100 nm resolution.
五十多年来，晶体学一直是研究蛋白质高分辨率结构的常规技术。大分子结构测定中的一个主要瓶颈是获得尺寸和质量适合单晶X射线晶体学实验的晶体。许多具有挑战性的蛋白质要么无法长成晶体，要么无法长成适合使用传统X射线晶体学获得高分辨率结构的尺寸的晶体。对于较小的晶体，它们可用于接种以获得更大的晶体，或用于串行晶体学和电子衍射以获得结构。对于这两个目的，一个限制步骤是对亚微米尺寸的此类小晶体进行非侵入性成像，并筛选此类晶体占主导的条件。在这里，我们使用基于阴极发光（CL）的纳米显微镜对蛋白质纳米晶体进行成像。我们表明，基于CL的纳米显微镜可以对微米和亚微米尺寸的晶体进行非侵入性成像。这里展示的结果证明了以低于100纳米的分辨率对蛋白质晶体进行非侵入性成像的可行性。
