Immobilization of Proteins in Silica Gel: Biochemical and Biophysical Properties

Abstract

Abstract:The development of silica-based sol-gel techniques compatible with the retention of protein structure and function started more than 20 years ago, mainly for the design of biotechnological devices or biomedical applications. Silica gels are optically transparent, exhibit good mechanical stability, are manufactured with different geometries, and are easily separated from the reaction media. Biomolecules encapsulated in silica gel normally retain their structural and functional properties, are stabilized with respect to chemical and physical insults, and can sometimes exhibit enhanced activity in comparison to the soluble form. This review briefly describes the chemistry of protein encapsulation within the pores of a silica gel three-dimensional network, the mechanism of interaction between the protein and the gel matrix, and its effects on protein structure, function, stability and dynamics. The main applications in the field of biosensor design are described. Special emphasis is devoted to silica gel encapsulation as a tool to selectively stabilize subsets of protein conformations for biochemical and biophysical studies, an application where silica-based encapsulation demonstrated superior performance with respect to other immobilization techniques.

More than a Confinement: “Soft” and “Hard” Enzyme Entrapment Modulates Biological Catalyst Function

“…A low degree of condensation, which in the case of silica gels corresponds to a high amount of free hydroxyl groups, high ability of water binding, and large pores, yields looser matrices; a high degree of condensation yields more rigid systems with smaller pores. This is the result of a different average numbers of covalent bonds per molecular unit, which generates a discontinuity in the behavior at the molecular scale: the rigidity increases stepwise and is defined at the moment of the gel formation, and any modulation arises from the presence of a mixed population of pores with different diameters or hydration [13].…”

“…For the same structural reasons (prevailing covalent bonds and low flexibility), in silica gel or other more structured matrices (e.g., zeolites), the confining environment is geometrically characterized: the dimensions and shapes of the pores that form in the rigid scaffold determine the confining ability, the type of macromolecules that can be embedded, and the properties of the "solvent" layer trapped within the pores [13]. The larger flexibility in saccharide confinement makes porosity not the primary measure of the confining ability: polysaccharide matrices with defined porosity do exist, but the macromolecular scaffold is in any case less rigid and allows a certain degree of adaptability, allowing also the hosting of macromolecules with dimensions or shapes not properly matching those of the pores.…”

Immobilization of Allantoinase for the Development of an Optical Biosensor of Oxidative Stress States

“…puuE was encapsulated in a silica matrix by the sol-gel method [24,25]. Tetramethylorthosilicate (TMOS), water and a 40 mM HCl solution were mixed and sonicated for 20 min (divided in 5-min on and 1-min off cycles), then an equal volume of 10 mM phosphate, pH 6.0, was added and the solution was fluxed with humidified nitrogen for 40 min.…”

“…The relevance of a biosensor resides in the selectivity and sensitivity in target quantification and reusability, determining its applicability as a tool for routine clinical practice. With this purpose, we entrapped puuE in a wet nanoporous silica gel matrix, a strategy largely exploited in enzyme immobilization because of its several advantages, among which the easy and flexible chemistry, with mild chemical and physical entrapment conditions compatible with protein stability, the optical transparency allowing signal detection with conventional spectroscopic techniques, the ease of separation of the active material from the reaction solution [24][25][26][27][28][29][30][31].…”

Transfer and Reactivity of Hydrogen Sulfide with Immobilized Hemeproteins in Polymeric Matrix

“…It is well-known that encapsulation of biomolecule, such as proteins [1] [2], enzymes [3], antibodies and cells [1], using sol-gels method is widely considered for research in the development of biosensor [4] [5] [6], tissue engineering [7], and drug delivery [8] [9]. This technique involves the inclusion of a biomolecule in an inorganic matrix where mobility is restricted but allows migration of particular analytes through the gel lattice [10] [11].…”