Search

Protein crystallization occurs when the solution conditions are suitable for nucleation and growth of the crystals. pH, as one of the most important parameters that governs the protein crystallization process, can affect the conformation, activity, electrostatic interactions and solubility of protein in the solution. Hence, manipulating the pH is an important path to success in crystallizing proteins. In this paper, we show that varying pH levels over the incubation time during the crystallization process can help increase the chance of obtaining protein crystals. The process of varying the pH can be considered as one way to automatically search for a suitable pH for successful crystallization. This discovery can guide us to develop a new screening strategy in which varying the pH against the incubation time is the major process used during the crystallization screening. Our screening experiments verified that varying the pH is indeed a promising method to achieve efficient crystallization screening of a protein. [ABSTRACT FROM AUTHOR]

Environments with varying temperatures have been shown to beneficially increase the probability of obtaining protein crystals. Therefore, a cycling temperature strategy (CTS) has been proposed for protein crystallization screening. During the practical application of this strategy, it is necessary to know the effective temperature range that promotes crystallization to design a suitable temperature program. In this paper, the effects of different temperature ranges on lysozyme crystallization (or more specifically, nucleation) were investigated. The results show that a small periodic variation in the temperature range of as little as 0.4 K can have a significant effect on the crystallization success rate under some crystallization concentration conditions, confirming that crystallization of lysozyme is very sensitive to temperature variation. Because practical protein crystallization is always performed in an environment with slight temperature variations, the sensitivity of protein crystallization to temperature may provide an explanation for the poor reproducibility of protein crystallization. Further investigation of the CTS on lysozyme crystallization showed that a cycling temperature strategy exerts an effect on protein crystallization by altering the supersaturation caused by changes in temperature. [ABSTRACT FROM AUTHOR]

The rapid advance in superconducting magnet technology enables more and more applications for the use of high magnetic fields in scientific researches and industrial manufacturing. These applications include material processing, separation, chemical reaction, nuclear fusion, high energy physics, and many more. Generally, a superconducting magnet provides both homogeneous and inhomogeneous magnetic fields simultaneously, and both can affect the samples in the field so that the magnetic field can be utilized for various purposes. A homogeneous or inhomogeneous magnetic field will exert a torque on suspending particles in a solution if the particles have anisotropic magnetic susceptibility, which will further influence the properties of the solution; in an inhomogeneous magnetic field, a repulsive force will act on a diamagnetic solution so that the levels of apparent or effective gravity of the solution can be tuned in a vertical magnetic field. These effects can be utilized to govern the physical and chemical processes in solution like crystallization. In recent years, high magnetic fields have been applied in protein crystallization. It was found that a magnetic field can align the crystals along the field direction, decrease the diffusivity of macromolecules in the solution, and increase the viscosity of the solution; a suitable inhomogeneous magnetic field can damp the natural convection substantially, which resembles the case in a space environment. Both homogeneous and inhomogeneous magnetic fields have been found to improve the quality of some protein crystals. These discoveries showed that the researches on protein crystallization in high magnetic field is potentially valuable, because obtaining high quality protein crystals is important for 3-dimensional structure determination of proteins using X ray crystallography. This paper will review the background and more recent progress and discuss the future perspectives in this research field. [ABSTRACT FROM AUTHOR]

It is well known that the crystallization of proteins is strongly dependent on the crystallization conditions, which are sometimes very sensitive to environmental disturbances. Parameters such as the concentration of precipitants or protein, pH, temperature and many others are known to affect the probability of crystallization, and the task of crystallizing a new protein often involves a trial-and-error test using numerous combinations of crystallization conditions. These crystallization parameters, such as the concentration of either the protein or the precipitant, are important because they directly affect the driving force of crystallization: the supersaturation of the solution. Although it is common sense that the concentration can affect the crystallization process, the sensitivity of the crystallization process to variations in the concentration has seldom been addressed. Owing to the difficulty of directly preparing solutions with very small concentration variations, it is hard to carry out an investigation of their effect on the crystallization process. In this paper, a simple but novel method for studying the effect of minute concentration variations on the success rate of protein crystallization is presented. By evaporating the crystallization droplet, a fine concentration gradient could be created. With this fine-tuned concentration gradient, it was possible to observe the effects of minute variations in the concentration or supersaturation on the crystallization. A very minor change in concentration (as low as 0.13% of the initial concentration, i.e. 0.026 mg ml−1 for lysozyme and 0.052 mg ml−1 for NaCl in the current study) or a very minor change in supersaturation (as small as 0.018) could cause a clear difference in the crystallization success rate, indicating that the crystallization of proteins is very sensitive to the concentration level. Such sensitive behaviour may be one reason for the poor reproducibility of protein crystallization. [ABSTRACT FROM AUTHOR]

• HD method can obtain more crystallization hits of "several diffraction spots". • HD method was beneficial for the quality of protein crystals. • HD method in CDM plate possess larger number of screening crystallization hits than that of T plate. Obtaining good quality protein crystals is still desirable for high resolution structural determination of the proteins using crystallography. The crystal quality is affected by the growth environment and growth methods. As frequently used methods in practical protein crystallization, sitting and hanging-drop methods are seldom compared in terms of crystal quality in the literatures. We performed a systematic comparison on the quality of the protein crystals grown in sitting and hanging-drop methods using different crystallization plates: cross-diffusion microbatch plate (CDM SD: CDM plate, sitting drop method; CDM HD: CDM plate, hanging drop method), traditional sitting and hanging-drop vapor diffusion plates (T SD: traditional plate, sitting drop method; and T HD: traditional plate, hanging drop method). Five different proteins, proteinase K (prk), lysozyme (lys), concanavalin A VI (con), catalase (cat) and α-Chymotrypsinogen A II (chy), were used. The crystal quality was compared in terms of the resolution limit, mosaicity and Wilson plot B factor. It was found that the crystals grown in CDM plate using the hanging drop method (CDM HD) exhibited the best morphology and the best crystal quality. X-ray diffraction tests showed that the CDM plate using hanging drop method is indeed a practical and useful method for obtaining high-quality protein crystals. [ABSTRACT FROM AUTHOR]

Surface treatment by oxidizing the crystallization plates can significantly promote protein crystallization and requires no change to routine screening protocols; therefore, it is potentially useful for practical protein crystallization screening. However, experiments have shown that the amount of oxidants and the treatment process need to be optimized to achieve effective results. Searching for the suitable amount of oxidants, temperature and processing time for surface treatment of the plate will increase the workload and decrease the efficiency of the screening process. To solve this problem, a series of trials to determine suitable surface treatment conditions were conducted. Based on these experiments, not only was the most suitable processing condition for the optimal protein crystallization screening hits discovered but also the relationship between the treatment process and the protein crystallization screening hits was explored. With these results, the surface treatment of protein crystallization plates became easier and more effective, allowing the oxidizing surface treatment method to be applied on plates for use in routine protein crystallization screening. [ABSTRACT FROM AUTHOR]