Document Type

Article

Publication Date

1-1-2008

Publication Title

Biophysical Journal

Volume

95

Issue Number

8

First Page

L51

Last Page

L53

DOI

10.1529/biophysj.108.142182

Disciplines

Biochemistry | Other Chemistry

Abstract

The secondary structures of two proteins were examined by circular dichroism spectroscopy after adsorption onto a series of organically modified silica glasses. The glasses were prepared by the sol-gel technique and were varied in hydrophobicity by incorporation of 5% methyl, propyl, trifluoropropyl, or n-hexyl silane. Both cytochrome c and apomyoglobin were found to lose secondary structure after adsorption onto the modified glasses. In the case of apomyoglobin, the α-helical content of the adsorbed protein ranged from 21% to 28%, well below the 62% helix found in solution. In contrast, these same glasses led to a striking increase in apomyoglobin structure when the protein was encapsulated within the pores during sol-gel processing: the helical content of apomyoglobin increased with increasing hydrophobicity from 18% in an unmodified glass to 67% in a 5% hexyl-modified glass. We propose that proteins preferentially adsorb onto unmodified regions of the silica surface, whereas encapsulated proteins are more susceptible to changes in surface hydration due to the proximity of the alkyl chain groups. Adsorption of proteins onto solid surfaces is an important issue for many fields of research, including the design of biomedical devices, biocatalyst and biosensor development, food manufacturing, and protein separation techniques (1). Adsorption phenomena are governed by many factors, including the charge and polarity of the interacting surfaces, the ease of desolvation, and the structural stability of the protein (2). Porous silica materials produced by the sol-gel technique provide an interesting system for testing the effects of surface chemistry on protein adsorption. Sol-gel-derived silica glasses are obtained by hydrolysis of a tetraalkoxysilane (typically tetramethyoxysilane (TMOS) or tetraethoxysilane) followed by condensation to form a network of Si-O-Si linkages (3). The tetraalkoxysilane may be supplemented with other silane precursors, e.g., RSi(OCH3)3, where R represents any nonhydrolyzable organic group, to incorporate new chemical functionality into the silica matrix. In this manner, glasses may be engineered to alter the charge, polarity, or hydrophobicity of the surface in contact with the solvent phase. Under many conditions, these glasses result in optically transparent materials that are amenable to spectroscopic techniques and therefore the study of protein structure after adsorption. Because of the inherently mild processing conditions involved, many investigators use the sol-gel technique to encapsulate biomolecules in porous glasses as a noncovalent means of immobilization; proteins may be added to the liquid sol before the condensation reaction that leads to formation of the matrix is initiated. This approach has led to several examples where the encapsulated protein maintains its native structure and biological activity 4 and 5. In the current study, silica glasses were made with TMOS and a series of RSi(OCH3)3 reagents (where R represents a methyl, propyl, trifluoropropyl, or n-hexyl group) to study adsorption as a function of increasing hydrophobicity. The choice of these functional groups was guided by previous work indicating that a low content of hydrophobic precursor can have a striking effect on the structure of an encapsulated protein 6 and 7. Because the addition of organic groups may alter the porosity of the glass in addition to the surface chemistry, it is important to check the physical properties of the bulk glass. As shown in Table 1, the 5% methyl, propyl, and hexyl glasses yield similar values for the specific surface area, pore volume, and average pore diameter. The control glass of 100% TMOS is characterized by larger pores and lower specific surface area than the alkyl-modified glasses, and the glass containing organic fluorine yields values intermediate between the two extremes

Comments

Copyright © 2008 Biophysical Society.

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