Properties of human serum albumin entrapped in sol-gel-derived silica bearing covalently tethered sugars

Silica derived from biocompatible silane precursors and containing covalently bound sugar moieties has recently been reported to be a much more biocompatible matrix for protein entrapment than any previously synthesized materials. To better understand the nature of these new materials, the steady-state and time-resolved fluorescence of human serum albumin (HSA) was used to examine the conformation, dynamics, accessibility, thermal stability, and degree of ligand binding after entrapment of the protein into sol-gel-processed glasses derived from either tetraethyl orthosilicate (TEOS) or diglycerylsilane (DGS), which in some cases contained covalently bound gluconamidylsilane (GLS) moieties. It was observed that the initial conformation, accessibility to external analytes, thermal stability, long-term stability, and degree of ligand binding to HSA were best in DGS-derived materials that contained covalently tethered GLS relative to unmodified DGS-derived materials, TEOS, or TEOS/GLS-derived materials. Measurement of protein rotational dynamics showed that entrapment led to an immediate loss of global motion in all materials. However, the restriction of motion was most dramatic in GLS-doped materials, suggesting preferential interactions of the protein with the sugar-coated surfaces. As aging proceeded, both protein dynamics and the degree of ligand binding decreased, with a gradual loss of segmental motion and a significant increase in local motion in the vicinity of the probe, consistent with unfolding and surface adsorption of the protein, leading to loss of function. Overall, our findings suggest that the use of a biocompatible precursor (DGS) and the addition of a covalently bound sugar both contribute to improved protein performance. However, of these two the use of a biocompatible precursor is the most important factor, and in such cases addition of sugars further improves protein performance. In contrast, the use of the sugar-based additive with a nonbiocompatible precursor such as TEOS imparted essentially no benefit, demonstrating the importance of biocompatible processing conditions.