Technique for High-Quality Protein Crystal Growth by Control of Subgrain Formation under an External Electric Field

Haruhiko Koizumi,Kenichi Kojima,Kozo Fujiwara,Jun Nozawa,Masaru Tachibana,Satoshi Uda

doi:10.3390/cryst6080095

Abstract

X-ray diffraction (XRD) rocking-curves were measured for tetragonal hen egg white (HEW) lysozyme crystals grown with and without application of an external electric field, and the crystal quality was assessed according to the full width at half-maximums (FWHMs) of each rocking-curve profile. The average FWHMs for tetragonal HEW lysozyme crystals grown with an external electric field at 1 MHz were smaller than those for crystals grown without, especially for the 12 12 0 reflection. The crystal homogeneity of the tetragonal HEW lysozyme crystals was also improved under application of an external electric field at 1 MHz, compared to that without. Improvement of the crystal quality of tetragonal HEW lysozyme crystals grown under an applied field is discussed with a focus on subgrain formation. In addition, the origin of subgrain misorientation is also discussed with respect to the incorporation of impurities into protein crystals.

Highlights

The structural analysis of protein molecules using X-ray diffraction (XRD) analysis is actively performed in order to achieve structure-guided drug design and controlled drug delivery
The improvement of the crystal quality of tetragonal hen egg white (HEW) lysozyme crystals as a model protein is demonstrated by applying an external electric field
We first focus on the average full width at half-maximums (FWHMs) obtained from crystals grown without an external electric field

Summary

Introduction

The structural analysis of protein molecules using X-ray diffraction (XRD) analysis is actively performed in order to achieve structure-guided drug design and controlled drug delivery. The improvement of the crystal quality of tetragonal hen egg white (HEW) lysozyme crystals as a model protein is demonstrated by applying an external electric field. We have succeeded in the control of both an increase and a decrease in the nucleation rate for tetragonal HEW lysozyme crystals, by applying an external electric field with different frequencies [49]. We have demonstrated that this crystallization technique can be adapted to the crystallization of porcine insulin [50] This is attributed to the difference in the magnitude of the electrostatic energy added to the chemical potentials between the liquid and solid phases. This article demonstrates that improvement of the quality of crystals grown under a 1 MHz applied field is achieved by a decrease in the misorientation between subgrains in the crystals. We discuss the improvement of the crystal quality under an applied field at

Experimental Procedure

Improvement of Crystal Quality under an Applied Electric Field

Control of Subgrain Formation under an Applied Electric Field