Gel growth of lysozyme crystals studied by small angle neutron scattering: case of agarose gel, a nucleation promotor

A crystallization technique for obtaining large protein crystals with increased mechanical stability using agarose gel combined with a stirring technique

“…By increasing the agarose gel concentration, the nucleation probability monotonically increased (see the rotating speed of 0 rpm). This tendency was consistent with the results reported previously [7], and was clearly an effect of the nucleation enhancement by agarose gel [7,30]. Increasing the rotating speed under the conditions of 0.5%, 1.0% and 2.0% agarose gel concentration decreased the number of crystals.…”

“…Among typical gels used for protein crystallization, agarose gels are the most employed hydrogels in protein crystallization because of its stability, easy for use and high familiarity for biochemists [6]. Crystallization using agarose gel has some advantages; the nucleation enhancement effect [7,30], the impurity filtering effect during crystal growth [31], the suppression of heat convective transport and crystal sedimentation [32], and the enhancement of crystal strength by incorporation of agarose gel fibers into protein crystals [6,8]. Especially the enhancement of crystal strength by agarose gel has great advantages for overcoming handling damage before X-ray diffraction studies or osmotic shock damage during ligand soaking experiments [8,9].…”

A supersaturation wave of protein crystallization

“…Certainly, under terrestrial conditions we can reduce convective transport by lowering the ratio between buoyancydriven convection and diffusion, for instance by decreasing the characteristic length of the system [22], i.e., using capillaries [23,24] or gels [25][26][27]. However, convective flow cannot be completely avoided inside X-ray capillaries of 0.2 mm inner diameter (the minimum crystal size for significant X-ray data collection of macromolecular crystals at conventional sources) and, with gels, the gel polymeric network may in some cases interact chemically with the protein molecules [28,29]. Nevertheless, the nowadays scarce flight opportunities, the existence of g-jitters and relevant values of residual accelerations on board the Shuttle [30] and the complex logistics associated with space experiments, makes it worth to develop research studies devoted to investigate in these on-ground analogs.…”

Enhancement of femtosecond laser-induced nucleation of protein in a gel solution

“…On the other hand, in the agarose gel condition, spontaneous nucleation occurred at lower ϳ 1.0. Numerous studies have reported such enhancement of nucleation by agarose, 19 and its mechanism was explained by promotion of the association among protein clusters. 19 However, we found that laser irradiation in the agarose gel condition realized the nucleation at the lowest ϳ 0.3, which is three times lower than that of the spontaneous nucleation.…”

“…Numerous studies have reported such enhancement of nucleation by agarose, 19 and its mechanism was explained by promotion of the association among protein clusters. 19 However, we found that laser irradiation in the agarose gel condition realized the nucleation at the lowest ϳ 0.3, which is three times lower than that of the spontaneous nucleation. This clearly indicates that the focused femtosecond laser irradiation in the presence of agarose gel enhances nucleation more than the laser or agarose alone.…”