Extraction and Characterization of Hydrophobin from Trichoderma reesei

Abstract:

Hydrophobins are a class of small proteins (7-15kDa) found in filamentous fungi and are among the most surface active proteins known today. Because of this they have received attention for different applications, e.g. for the food industry as an alternative in emulsions. The goal of this project was to culture and extract hydrophobins from Trichoderma reesei and characterize it. This was done from a freeze-dried culture of Trichoderma reesei, which was cultured on PDA-plates and in liquid medium with glucose as carbon source. Extraction was made by breaking the cells, mechanically and by sonication, and then by shaking a seperating funnel to create foam from the surface-active proteins. The foam was washed and freeze-dried and the total protein concentration of the freeze-dried substance was determined with Bradford assay and the hydrophpbin was characterized with SDS-PAGE. The culturing of the fungi was successful. The amount of foam created was, however, less than expected. The Bradford assay gave a total protein concentration of 7.5% in the freeze-dried substance, but the SDS-PAGE didn't give any results. The reason for this probably depends on the culturing and the extraction of the hydrophobin. T. reesei hydrophobin HFBI, expressed in glucose containing media, is bound to the mycelium of the fungi and the breaking of the mycelium might not have been enough to release all the protein, which also would explain the small amounts of foam. One way to improve this could be to grow the fungi on lactose instead. This will result in that T. reesei produces HFBII instead, which is mainly released to the surrounding. The conclusion of the project is that the method for culturing and extraction needs to be improved to obtain hydrophobin from T. reesei.