Methods for Obtaining Better Diffractive Protein Crystals: From Sample Evaluation to Space Crystallization

Abstract

In this paper, we present a summary on how to obtain protein crystals from which better diffraction images can be produced. In particular, we describe, in detail, quality evaluation of the protein sample, the crystallization conditions and methods, flash-cooling protection of the crystal, and crystallization under a microgravity environment. Our approach to protein crystallization relies on a theoretical understanding of the mechanisms of crystal growth. They are useful not only for space experiments, but also for crystallization in the laboratory.

Through Diffusion Measurements of Molecules to a Numerical Model for Protein Crystallization in Viscous Polyethylene Glycol Solution

“…The dotted lines in Figure 5 are the linear regression of the dots, showing that k 5 , k 6 , k 7 and k 8 are 0.6804, 0.1450, 3.819 × 10 −4 and 1.1456, respectively. Then, the anomalous index, α in the Equation (7), is as follows, using the estimated values: Finally, using Equation (10), we are able to estimate the anomalous index α, so that τ D of a solute molecule in an arbitrary PEG solvent can be estimated by the following equation.…”

“…Thus, automated microfluidic systems that produce crystals using a counter-diffusion technique have been developed to save the protein sample and obtain good crystals [8]. Counter-diffusion (also known as liquid-liquid diffusion) is a common crystallization method, in which the protein and precipitants are loaded on opposite sides of a tubing chamber and gradually mixed through their diffusion [3,9,10]. In most cases, the protein is loaded into the capillary, the end of which is sealed with a gel plug to keep the protein from flowing out and allow the precipitant to diffuse in from the reservoir.…”

Procedure of the overexpression, purification and crystallization of BLEG-1, a bifunctional and evolutionary divergent B3 metallo-β-lactamase, for structure-function studies

“…The obtainment of protein crystals of good quality (i.e. single crystal with medium-to-large size and of appropriate shape) for diffraction is often desired, as this would increase the probability of obtaining diffraction data and eventually a protein structure of higher quality [13] . For this purpose, it is necessary to produce protein sample at high concentration, homogeneity, solubility and stability.…”

Enhancement of Continuous Crystallization of Lysozyme through Ultrasound

“…As a traditional protein crystallization method, salting-out crystallization usually produces crystals with a wide particle size distribution, , which seriously affects the quality of the final products. Besides, commonly used batch operation in protein crystallization also has some disadvantages, such as batch-to-batch quality variation, time and labor-consuming, and high cost. ,, With the increasing demand for high quality and efficient production of crystalline materials, continuous crystallization has attracted extensive attention from the pharmaceutical industry and academia.…”

“…As a traditional protein crystallization method, salting-out crystallization usually produces crystals with a wide particle size distribution, 17,18 which seriously affects the quality of the final products. Besides, commonly used batch operation in protein crystallization also has some disadvantages, such as batch-to-batch quality variation, time and labor-consuming, and high cost.…”