Characterization of hydrophobin SC3 interfacial self-assembly

Jeffrey Shawn Goodwin

Abstract

Amphipathic fungal proteins called hydrophobins are able to self-assemble into insoluble, supramolecular structures at hydrophobic/hydrophilic interfaces. This property allows the hydrophobin SC3 to assemble onto hydrophobic liquid microdispersed in an aqueous solution. As a result of this action, SC3 is able to sequester hydrophobe into protein encapsulated vesicle-like structures. The protein coat surrounding these vesicles was identified using epifluorescence microscopy by fluorescently labeling SC3 via antibodies against SC3. Likewise, the content of the vesicles was characterized as hydrophobic using the fluorescent probe Nile Red, while the exterior was determined to be hydrophilic using the fluorescent probe calcein. By investigating the interfacial self-assembly of the protein, evidence was provided in this study that supports two possible applications: water remediation and drug delivery. Little is known about the molecular mechanism and underlying protein conformational changes occurring during the self-assembly of hydrophobins. In this investigation, it was found that the self-assembled form of SC3 resembles amyloid β-fibrils. Nevertheless, SC3 self-assembly is promoted by the presence of an interface, while the amyloid-β protein assembly is nucleated by preassembled protein fibrils. Once the interface is coated with self-assembled SC3, further loss of soluble protein appears to occur from random aggregation. When SC3 self-assembles, the molecules form a stacked β-sheet structure and present a more hydrophobic surface to the aqueous solution, suggesting that a structural change is occurring as the protein self-assembles.