Relaxation of a large amount of Local Strain in Protein Crystals by using Seed Crystals

H. Koizumi¹, S. Uda¹, M. Tachibana², K. Tsukamoto³, K. Kojima⁴, J. Nozawa¹

¹Institute for Materials Research, Tohoku University, 2-1-1, Katahira, Aoba-ku, Sendai, 980-8577, Japan

²Graduate School of Nanobioscience, Yokohama City University, 22-2 Seto, Kanazawa-ku, Yokohama, 236-0027, Japan

³Graduate School of Engineering, Osaka University, 2-1 Yamadaoka, Suita, Osaka 565-0871, Japan

⁴Department of Education, Yokohama Soei University, 1 Miho-tyou, Midori-ku, Yokohama, 226-0015, Japan

h_koizumi@imr.tohoku.ac.jp

In order to allow both structure-guided drug design and controlled drug delivery, it is important to determine the 3D structures of protein molecules. At present, these structures are primarily determined using X-ray and neutron diffraction analysis, and therefore high-quality single crystals of proteins are required, even though these are quite difficult to obtain. For neutron diffraction analysis, in particular, large protein crystals are needed to detect many diffraction spots because of the weakness of the brightness of neutron beam. However, it is quite difficult to grow large protein crystals. This could be attributed to a large amount of local strain in the crystal. Therefore, the establishment of a crystallization technique to obtain large high-quality single crystals of proteins is desired.

Recently, we have revealed that imperfections in tetragonal hen egg white (HEW) lysozyme crystals are predominantly caused by local strain, < ε >, and misorientation between subgrains, |θ ‒ φ|, in the crystal, by analyzing the full width at half-maximum (FWHM) of the rocking curves [1] (see Table 1). Moreover, we have demonstrated that crystal quality of protein crystals can be improved by application of an external electric field [2, 3], which is achieved by a decrease in the misorientation between subgrains in the crystal [1]. That is, the misorientation between subgrains can be controlled by applying an external electric field. However, there is a large amount of local strain in protein crystals. In order to achieve large high-quality single crystals of proteins, therefore, not only control of the misorientation between subgrains, but also relaxation of a large amount of local strain in protein crystals must be preformed. In this presentation, we indicate that the crystallization technique using seed crystals is useful to release a large amount of local strain in protein crystals. This crystallization technique has hidden potential to grow large high-quality single crystals of proteins which is required for neutron diffraction analysis.

Table 1. Misorientation between subgrains, |θ ‒ φ|, and local strain, < ε >, for tetragonal HEW lysozyme crystals.

	Tetragonal HEW lysozyme
Misorientation	0.0031°
Local strain	137 µε

1. H. Koizumi, S. Uda, K. Fujiwara, M. Tachibana, K. Kojima, and J. Nozawa, Cryst. Growth Des. 14, (2014), 5662.

2. H. Koizumi, S. Uda, K. Fujiwara, M. Tachibana, K. Kojima and J. Nozawa, J. Appl. Cryst., 46, (2013), 25.

3. H. Koizumi, S. Uda, K. Fujiwara, M. Tachibana, K. Kojima and J. Nozawa, AIP Conference Proceedings, 1618, (2014), 265.