Fluoro-Silane as a Functional Monomer for Protein Conformational Imprinting

Silica gels capable of inducing defined conformational changes in proteins were synthesized by polymerization of either 3,3,3-trifluoropropylmethoxysilane (3F) or (3-heptafluoroisopropoxy)propalethoxysilane (7F) with tetraethylorthosilicate (TEOS). Bovine serum albumin (BSA) and fluorescein isothiocyanate conjugated BSA (FITC-BSA) were encapsulated in the gels. The hydrolyzed TEOS sol mixing with 0.01 M pH 8 hepes buffer at a ratio of 1:3 (sol:buffer) was selected as the optimal condition for rapid synthesis of rigid and optically transparent gels. These 1:3 gels doped with 10% 3F (3F/TEOS, mol%) or with 5% 7F (7F/TEOS, mol%) were as transparent as pure TEOS gels, whereas 10% 7F modification resulted in a turbid gel containing micron-sized 7F aggregates that excluded encapsulated FITC-BSA. In contrast, FITC-BSA was homogenously dispersed in the pure TEOS and 10% 3F modified TEOS gels. Water contact angle increased from 19.9o ± 2.6o on pure TEOS gel to 85.8o ± 3.4o on 5% 3F-modified gels and to 105.4o ± 1.1o on the 5% 7F gels. However, 10% 7F modification reversed the hydrophobicity trend by reducing the contact angel to 55.2o ± 4.2o. The fluorospectroscopic assays on the fluoro-silane-BSA interactions showed that either hydrolyzed 3F monomers or 3F copolymerized up to 10% in TEOS induced a concentration-dependent fluorescence reduction for 1-anilinonaphthalene-8-sulfonic acid (1,8-ANS), a lipophilic probe that has high affinity with the hydrophobic pockets of 69 proteins. The 3F-induced probe emission reduction may be interpreted as strong BSA-3F interactions excluding ANS-protein interactions. In contrast, hydrolyzed 7F increased the probe fluorescence, while 5% 7F in TEOS gel quenched the emission, suggesting that solution 7F facilitates ANS binding to BSA by exposing hydrophobic pockets on the protein; however, upon condensation into a TEOS gel, this effect is lost, possibly due to steric constraints in the gel. These findings demonstrate the advantages of 3F over 7F as a functional monomer for TEOS gel modification in terms of retaining gel transparency and increasing hydrophobicity (and consequently stability against capillary-induced collapse) while maintaining a uniform distribution of encapsulated protein. Encapsulated protein appears to be stabilized by 3F binding to hydrophobic domains or clefts on the protein as evidenced by exclusion of the lipophilic probe, 1,8-ANS.