Title

Purification and Characterization of a New Class of Amphipathic Biopolymers Capable of Self-Assembly From Aqueous Media

Date of Award

2000

Degree Type

Dissertation

Degree Name

Doctor of Philosophy (PhD)

Department

Polymers and High Performance Materials

First Advisor

Charles L. McCormick

Advisor Department

Polymers and High Performance Materials

Abstract

Sc3p is the major hydrophobin produced and excreted by the wood-rotting fungus Schizophyllum commune when grown as a monokaryon in culture. In addition to Sc3p, several lower molecular weight proteins of unknown function and a high molecular weight polysaccharide, schizophyllan, are present in this fungal culture medium. Schizophyllan and hydrophobin, as produced in the culture supernatant, work in concert to ultimately produce stable films on hydrophilic substrates. A procedure was developed that facilitates the purification of the Sc3p hydrophobin without having to induce protein aggregation in solution and thus eliminates the necessity of previously utilized organic solvent treatment. This procedure allows a systematic evaluation of the importance of the hydrophobin and schizophyllan components in solution stabilization and film formation from a conformational state more representative of that found during fungal production. These results indicate that schizophyllan stabilizes small hydrophobin multimers in aqueous solution. In addition, schizophyllan is necessary for subsequent hydrophobin assembly into thin films on hydrophilic surfaces. By contrast, hydrophobin alone is sufficient for thin film assembly to a hydrophobic surface; however, hydrophobin aggregation at high protein concentration in the absence of schizophyllan may limit the efficiency of film formation. These data suggest a model in which the polysaccharide acts as a macromolecular stabilizer for hydrophobin in solution, thereby preventing nonproductive self-aggregation. Schizophyllan may also allow lateral reorganization of the hydrophobin upon interaction with a hydrophilic surface such as the fungal cell wall, thereby providing a hydrophobic coating for protection and/or attachment to a host organism.