Background: EPL1 belongs to the cerato-platanin protein family found exclusively in fungi and associated with fungus-host interactions. Results: EPL1 self-assembles at air/water interfaces, increases the polarity of surfaces and solutions, and binds to chitin. Conclusion:The reported properties for EPL1 show that cerato-platanin proteins are clearly different from hydrophobins. Significance: This study reports several novel properties for cerato-platanin proteins.

Five bituminous samples were carefully studied by confocal laser scanning microscopy using 488 nm excitation radiation and observing 500-530 nm of emission. The images revealed the microstructure of bitumen. The influence of the admixture of mineral aggregates concerning the microstructure was tested. For the minerals, no significant influence was found. For understanding the origin of fluorescent signals, the samples were separated into asphaltenes and maltenes and analyzed with fluorescence spectroscopy. Although former works have assumed the origin of fluorescent emissions in bitumen to be found in the asphaltene fraction, the asphaltenes produce little to no emissions, but the maltenes exhibit strong fluorescence in the observed spectral region. For deeper insight, fractionation of the bitumina into the SARA fractions by chromatographic column separation was necessary. The fluorescence spectra of these fractions were analyzed and revealed the aromatics and resin phases to be the only components capable of sufficiently intense fluorescent emission. This is a strong argument for a complex internal microstructure consisting of a mantle of aromatics surrounding an inner core.

Cerato-platanin proteins (CPPs) and hydrophobins are two classes of small, secreted proteins that are exclusively found in fungi. CPPs are known as chitin-binding proteins, and were recently also shown to form protein layers at air/water interfaces, but the features of these layers were not investigated on the molecular level yet. In this study, by means of atomic force microscopy (AFM), EPL1, a member of the CPP family was shown to form highly ordered monolayers at a hydrophobic surface/liquid-interface. Furthermore, two new hydrophobins were analysed, and the influence of EPL1 on hydrophobin layers was studied in situ. Hydrophobins are amphiphilic proteins that are able to self-assemble at hydrophobic/hydrophilic interfaces, thereby inverting the polarity of the surface. This renders fungal growth structures such as spores water repellent. The combination of AFM data and wettability experiments led to the conclusion that in presence of both, hydrophobins and EPL1, a previously unknown hybrid layer is formed. This mixed protein layer is on one hand not inverting but enhancing the hydrophobicity of HOPG (highly oriented pyrolytic graphite), typical for EPL1, and on the other hand, it is stable and water insoluble, which is reminiscent of hydrophobin layers.