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. 2024 Nov 22;57(Pt 6):1907-1912.
doi: 10.1107/S1600576724010124. eCollection 2024 Dec 1.

Non-invasive nanoscale imaging of protein micro- and nanocrystals for screening crystallization conditions

Krishna Prasad Khakurel  1 , Kei Hosomi  2 , Wataru Inami  2 , Kawata Yoshimasa  2
Affiliations

Non-invasive nanoscale imaging of protein micro- and nanocrystals for screening crystallization conditions

Krishna Prasad Khakurel et al. J Appl Crystallogr. .

Abstract

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.

Keywords: cathodoluminescence; nanocrystals; nanoscopy; protein crystal screening.

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Figures

Figure 1
Figure 1
A schematic of the CL-EXA microscope discussed in this article.
Figure 2
Figure 2
(a) Scanning electron micrograph image of the Au NPs. (b) The CL-EXA image of the Au NPs at low magnification and (c) at high magnification. (d) A line plot across the red line shown in (c).
Figure 3
Figure 3
Monte Carlo simulation to estimate the penetration of an electron though the ZnO film to the protein crystal.
Figure 4
Figure 4
(a) CL-EXA images of the protein crystal. (b) Optical micrograph of the edge of the protein crystal. (c) The CL-EXA image of the edge of the protein crystal. (d) A line plot along the red line in (c), and (e) a line plot along the green line in (b).
Figure 5
Figure 5
(a), (d) CL-EXA images of a sub-micrometre protein crystal. (b), (e) FE-SEM images of the crystal shown in (a) and (d), respectively. (c), (f) Line plots along the red lines in (a) and (d), respectively.
See this image and copyright information in PMC

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