Interfacial rheology of globular and flexible proteins at the hexadecane/water interface: Comparison of shear and dilatation deformation

After long-time exposure, protein adsorption at fluid/fluid interfaces is documented to produce interfacial, gellike networks. Formation of this network apparently results from adsorption-induced conformational changes and subsequent interprotein aggregation at the interface. We utilize interfacial shear and dilatational rheology to probe the structure of a globular protein, lysozyme, and a disordered protein, beta-casein, and the kinetics of network formation at the hexadecane/water interface. For the first time, we present a detailed comparison of the interfacial shear and dilatational responses. For lysozyme, the shear moduli grow with interface age indicating a transition from fluidlike behavior at early times to network formation (solidlike behavior). Conversely, the interfacial shear moduli of beta-casein change very little with interface age; in addition, both G' and G" for beta-casein are an order of magnitude smaller than those of lysozyme. The strong protein intramolecular interactions that stabilize the native conformation of lysozyme act as kinetic barriers to conformational change and later become strong intermolecular interactions upon partial unfolding at the interface. Hence, interprotein linkages form (i.e., aggregation into an interfacial gel), resulting in the growth of G' with time. We find that the interfacial dilatational storage modulus, E', is comprised of a static response and a dynamic response. The static response corresponds to a change in the surface pressure upon interfacial-area change and is strain-rate independent. The dynamic contribution corresponds to rearrangement and reconfiguration of the protein molecules within the interface and is analogous to the shear storage response (i.e., a measure of the strength of interprotein linkages). The magnitudes of E' and G' for lysozyme and P-casein suggest that lysozyme initially adsorbs in a state similar to its native conformation. The native rigidity of the protein is linked to its kinetic stability at the interface. Globular lysozyme, once adsorbed, resists compression giving a high dilatational storage modulus. Contrastingly, native beta-casein lacks tertiary structure, resulting in a small interfacial dilatational storage modulus relative to lysozyme. With increasing interface age, the static modulus of beta-casein changes insignificantly, whereas it decreases substantially for lysozyme, indicating partial unfolding and loss of intrinsic rigidity. Upon unfolding, interprotein linkages form through hydrophobic peptide-peptide interactions. Correspondingly, G' and the recoverable dilatational storage modulus, deltaE', grow, signifying the onset of interfacial gelation.