Effects of Adsorption to Aluminum Salt Adjuvants on the Structure and Stability of Model Protein Antigens

Abstract

The effect of adsorption onto aluminum salt adjuvants on the structure and stability of three model protein antigens was studied using fluorescence and Fourier transform infrared spectroscopies, as well as isothermal titration and differential scanning calorimetric techniques. Lysozyme was preferentially adsorbed to aluminum phosphate (Adju-Phos®), whereas ovalbumin and bovine serum albumin were better adsorbed to aluminum hydroxide (Alhydrogel®). A linearized Langmuir adsorption isotherm was used to obtain information regarding the binding interactions between proteins and adjuvants. Binding energetics and stoichiometry data obtained from isothermal titration calorimetry measurements were complex. Based on the spectroscopic and differential scanning calorimetry studies, the structure of all three proteins, when adsorbed to the surface of an aluminum salt, was altered in such a way as to render the proteins less thermally stable. Besides the pharmaceutical significance of this destabilization, we consider the possibility that this phenomenon may facilitate the presentation of antigens and thus contribute to the adjuvant activity of the aluminum salts.

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“…7). It has been reported that interactions of protein antigens with the highly charged aluminum salts may destabilize their structures (81). Consistent with this finding, MPERp in contact with alum underwent significant aggregation, as denoted by the IR absorption at 1620 cm Ϫ1 (Fig.…”

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“…This probably reflects the particular robust structure of these VLPs, as adsorption to aluminum can result in more drastic changes in thermal stability with other antigens. 28 Nevertheless, a minor contribution could be detected and was attributed to a pre-transition at slightly lower temperature, requiring less energy, while the main transition Tm remained unaffected. Such remarkable conformational robustness can account, at least in part, for the excellent long-term stability of the vaccine.…”

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“…In theory, conformational changes may also occur when a vaccine is adsorbed to the adjuvant that affect its antigenicity. Jones et al [16] found significant changes in the shape of ATR-FTIR spectra for bovine serum albumin, lysozyme and ovalbumin upon adsorption to aluminum adjuvants. Those authors concluded that a disruption of antigen conformation caused by adsorption destabilized the proteins' structures, and proposed that this phenomenon may itself contribute to the adjuvant effect.…”

“…To yield adequate signal, the method we employed required higher protein concentrations than were present in the formulation for vaccination (0.7-0.9 mg/mL versus 0.2 mg/mL), but much less than is typically used in transmission FTIR (>10 mg/mL), or HATR on protein/adjuvant mixtures without deuteration (≥2 mg/mL) [16]. We used protein concentrations as low as possible in order to prevent the appearance of artifacts due to self-association at high concentration.…”