Protein Crystallization in Ionic-Liquid Hydrogel Composite Membranes

Abstract

Protein crystallization is a powerful purification tool. It is the first step for crystallographic structural investigations, and can be preparatory for biotechnological applications. However, crystallizing proteins is challenging and methods to control the crystallization process are needed. Ionic-liquid hydrogel composite membranes (IL-HCMs) have been used here as material capable of supporting protein crystallization and hosting grown crystals. We found that IL-HCMs affect the selection mechanism of glucose isomerase (GI) polymorphs and make GI crystals grow completely immersed into the hydrogel layer. X-ray diffraction studies show that IL ions do not bind to the protein, likely because IL molecules are constrained in the polymeric framework. Our GI crystal structures have been compared with many existing GI crystal structures using multivariate analysis tools, allowing a comprehensive overview of factors determining structural similarities, i.e., temperature variations and external stresses exerted during or after crystal growth, such as dehydration or presence of hydrogel of a different nature. GI crystals grown on IL-HCM fit perfectly in this framework, showing typical features induced by external forces. Overall, protein crystallization by IL-HCMs show potential for biotechnological applications, as it could constitute a natural means for containing crystallized enzymes in working conditions.

Crystal Growth in Gels from the Mechanisms of Crystal Growth to Control of Polymorphism: New Trends on Theoretical and Experimental Aspects

“…Crystals grow completely immersed in the hydrogel layer, probably by the specific interaction of proteins with the ionic liquids. This growth has shown to have an influence on space group and polymorphism [99].…”

Preparation of Cellulose Nanocrystal-Reinforced Physical Hydrogels for Actuator Application

“…In the last few decades, researchers have focused on synthesizing nanocomposite hydrogels for tissue engineering, drug delivery, antimicrobial, bioactive electrode coatings, actuators, and sensors [1][2][3][4]. Nanocomposite hydrogels are advantageous with softness, biocompatibility, multi-functionality, and adaptability.…”

Application of Polymers as a Tool in Crystallization—A Review

“…Poly(sodium 4-styrenesulfonate) ZnO [28] Poly(Citric Acid-co-Glycerol) CaCO 3 [99] Itaconic acid-2-acrylamido-2-methylpropanesulfonic acid allyl polyoxyethylene ether copolymer CaCO 3 [100] Organic polymer Struvit [101] Regiorandom pentacene-bithiophene, poly(3-hexylthiophene) 6,13-bis(triisopropylsilylethynyl)pentacene [102] Poly(ethylene glycol) Perovskite [104] Poly(3-hexylthiophene), poly(methyl methacrylate) Bis(triisopropylsilylethynyl) pentacene [108] Maleic acid and sodium ρ-styrenesulfonate copolymer CaSO 4 •2H 2 O [109] Poly(N-vinyl pyrrolidone) copolymers H 2 O [113] Composite membranes Glucose Isomerase [117] In conclusion, the authors would like to emphasize that the branch of science tackling complicated interactions between crystallizing solutions and polymer additives becomes currently more and more important from both theoretical and practical points of view and therefore is a promising area for further research.…”

“…Profio et al demonstrated the fabrication of hydrogel membranes displaying controlled chemical composition and nanostructure in the creation of crystals with enhanced diffraction properties at a lower protein concentration than the conventional technique [ 116 ]. Belviso et al used ionic-liquid hydrogel composite membranes as material for supporting protein crystallization [ 117 ]. They proved that protein crystallization by ionic-liquid hydrogel composite membranes had the potential for biotechnological applications and could contain crystallized enzymes in working conditions.…”