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Acta Crystallogr D Struct Biol. 2016 May;72(Pt 5):603-15. doi: 10.1107/S2059798316001546. Epub 2016 Apr 26.

Transmission electron microscopy for the evaluation and optimization of crystal growth.

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Department of Structural Biology, University of Pittsburgh School of Medicine, 3501 Fifth Avenue, Pittsburgh, PA 15260, USA.
Department of Pharmacology and Chemical Biology, University of Pittsburgh School of Medicine, M240 Scaife Hall, 3550 Terrace Street, Pittsburgh, PA 15261, USA.
JAN Scientific Inc., 4726 11th Avenue NE, Suite 101, Seattle, WA 98105, USA.
School of Life Sciences, Arizona State University, PO Box 874501, Tempe, AZ 85287, USA.
Endocrine Unit, Massachusetts General Hospital and Harvard Medical School, Boston, MA 02114, USA.
Biosciences Division, Argonne National Laboratory, 9700 South Cass Ave, Lemont, IL 60439, USA.
Stanford Synchrotron Radiation Lightsource, SLAC National Accelerator Laboratory, 2575 Sand Hill Road, Menlo Park, CA 94025, USA.


The crystallization of protein samples remains the most significant challenge in structure determination by X-ray crystallography. Here, the effectiveness of transmission electron microscopy (TEM) analysis to aid in the crystallization of biological macromolecules is demonstrated. It was found that the presence of well ordered lattices with higher order Bragg spots, revealed by Fourier analysis of TEM images, is a good predictor of diffraction-quality crystals. Moreover, the use of TEM allowed (i) comparison of lattice quality among crystals from different conditions in crystallization screens; (ii) the detection of crystal pathologies that could contribute to poor X-ray diffraction, including crystal lattice defects, anisotropic diffraction and crystal contamination by heavy protein aggregates and nanocrystal nuclei; (iii) the qualitative estimation of crystal solvent content to explore the effect of lattice dehydration on diffraction and (iv) the selection of high-quality crystal fragments for microseeding experiments to generate reproducibly larger sized crystals. Applications to X-ray free-electron laser (XFEL) and micro-electron diffraction (microED) experiments are also discussed.


X-ray free-electron lasers; XFELs; crystal optimization; micro-electron diffraction; nanocrystallography; structural biology; transmission electron microscopy

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