Surface Properties of Protein-Polyelectrolyte Solutions. Impact of Polyelectrolyte Hydrophobicity.

The strong influence of an amphiphilic polyelectrolyte, poly(N,N-diallyl-N-hexyl-N-methylammonium chloride), on the surface properties of solutions of globular proteins (lysozyme, ß-lactoglobulin, bovine serum albumin, and green fluorescent protein) depends on the protein structure and allows elucidation of the contribution of hydrophobic interactions in the protein-polyelectrolyte complex formation at the liquid-gas interface. At the beginning of adsorption, the surface properties are determined by the unbound amphiphilic component, but the influence of the protein-polyelectrolyte complexes of high surface activity increases at the approach to equilibrium. The kinetic dependencies of the dilational dynamic surface elasticity with one or two local maxima give a possibility to distinguish clearly between different steps of the adsorption process and to trace the formation of the distal region of the adsorption layer. The conclusions from the surface rheological data are corroborated by ellipsometric and tensiometric results.

Dynamic Properties of Adsorption Layers of κ-Casein Fibrils.

The dynamic surface properties of native κ-casein solutions and aqueous dispersions of its fibrils differ significantly from the corresponding properties of the systems with globular proteins. The dependence of the dynamic surface elasticity of κ-casein solutions on surface pressure has a local maximum, indicating partial displacement of macromolecules from the proximal region of the surface layer to the distal one. This dependence becomes monotonic for fibril dispersions, similar to the results for dispersions of globular protein fibrils, but unlike the latter case, the surface elasticity close to the steady state reaches values that are approximately four times higher than the data for native protein solutions at the same concentrations.

Adsorption of Denaturated Lysozyme at the Air-Water Interface: Structure and Morphology.

The application of protein deuteration and high flux neutron reflectometry has allowed a comparison of the adsorption properties of lysozyme at the air-water interface from dilute solutions in the absence and presence of high concentrations of two strong denaturants: urea and guanidine hydrochloride (GuHCl). The surface excess and adsorption layer thickness were resolved and complemented by images of the mesoscopic lateral morphology from Brewster angle microscopy. It was revealed that the thickness of the adsorption layer in the absence of added denaturants is less than the short axial length of the lysozyme molecule, which indicates deformation of the globules at the interface. Two-dimensional elongated aggregates in the surface layer merge over time to form an extensive network at the approach to steady state. Addition of denaturants in the bulk results in an acceleration of adsorption and an increase of the adsorption layer thickness. These results are attributed to incomplete collapse of the globules in the bulk from the effects of the denaturants as a result of interactions between remote amino acid residues. Both effects may be connected to an increase of the effective total volume of macromolecules due to the changes of their tertiary structure, that is, the formation of molten globules under the influence of urea and the partial unfolding of globules under the influence of GuHCl. In the former case, the increase of globule hydrophobicity leads to cooperative aggregation in the surface layer during adsorption. Unlike in the case of solutions without denaturants, the surface aggregates are short and wormlike, their size does not change with time, and they do not merge to form an extensive network at the approach to steady state. To the best of our knowledge, these are the first observations of cooperative aggregation in lysozyme adsorption layers.

Spread Layers of Lysozyme Microgel at Liquid Surface.

The spread layers of lysozyme (LYS) microgel particles were studied by surface dilational rheology, infrared reflection-absorption spectra, Brewster angle microscopy, atomic force microscopy, and scanning electron microscopy. It is shown that the properties of LYS microgel layers differ significantly from those of ß-lactoglobulin (BLG) microgel layers. In the latter case, the spread protein layer is mainly a monolayer, and the interactions between particles lead to the increase in the dynamic surface elasticity by up to 140 mN/m. In contrast, the dynamic elasticity of the LYS microgel layer does not exceed the values for pure protein layers. The compression isotherms also do not exhibit specific features of the layer collapse that are characteristic for the layers of BLG aggregates. LYS aggregates form trough three-dimensional clusters directly during the spreading process, and protein spherulites do not spread further along the interface. As a result, the liquid surface contains large, almost empty regions and some patches of high local concentration of the microgel particles.

Dynamic Surface Properties of Mixed Dispersions of Silica Nanoparticles and Lysozyme.

The surface properties of mixed aqueous dispersions of lysozyme and silica nanoparticles were studied using surface-sensitive techniques in order to gain insight into the mechanism of the simultaneous adsorption of protein/nanoparticle complexes and free protein as well as the resulting layer morphologies. The properties were first monitored in situ during adsorption at the air/water interface using dilatational surface rheology, ellipsometry, and Brewster angle microscopy. Two main steps in the evolution of the surface properties were identified. First, the adsorption of complexes did not lead to significant deviations in the dynamic surface elasticity and dynamic surface pressure from those for a layer of adsorbed lysozyme globules. Second, through the gradual displacement of protein globules from the interfacial layer as a result of further complex adsorption, the layer became more dense with much higher dynamic surface elasticity (∼280 mN/m compared to ∼80 mN/m for a pure protein layer). These layers were shown to be fragile and could be easily broken into separate islands of irregular shape by a weak mechanical disturbance. The layer properties were then monitored following their transfer to solid substrates using atomic force microscopy and scanning electron microscopy. These layers were shown to consist of nanoparticles surrounded by a rough shell of protein globules, whereas some particles tended to form filamentous aggregates. This comprehensive study provides new mechanistic and morphological insight into the surface properties of a model protein/nanoparticle system, which is of fundamental interest in colloidal science and can be extended to systems of physiological relevance.