Struktur- und Funktionsanalysen von pilzlichen Zellwandproteinen der SUN- und CFEM-Proteinfamilien

The fungal kingdom is one of the most diverse lifeforms found on earth. Hereby the manifold number of different subspecies reflects their populated ecological niches. The fungal cell wall is a complex and highly functional organelle that allows adaptation to adverse environmental conditions like change in osmolarity. This 50 – 200 nm thick layer is formed by a basic set-up from chitin and b- 1,3-/ b-1,6-glucans and further species specific content. Within this heavily cross-linked network the b-1,3-glucans form triple helical structures that are then linked to the chitin moiety. This corecontent of b-glucans and chitin is primarily responsible for the robustness of the fungal cell wall. In the cross–linked space of the inner cell wall an enormous number of secreted proteins perform various roles. This work will focus on the two groups of SUN- and CFEM-proteins. SUN-proteins play a role during the cell wall remodelling process of b-1,3-glucans. SPR and HDX-MS experiments for the Cterminal SUN-domain of Saccharomyces cerevisiae showed active binding towards Laminarin, a b-1,3- glucan with additional b-1,6-glucose linkage [7:1]. Molecular dynamic simulations of ScSun4C with bound curdlan, an unlinked triple helical b-1,3-glucan, showed stable binding during the whole calculation of 100 ns. These binding results combined with structural information of the C-terminal SUN-domain lead to two hypothetical models. The SUN-domain as a novel, reversible b-glucan docking unit or as a triple helical b-glucan unwinding domain. This glucan helicase like behaviour unwinds hereby the triple helical form and passes one single b-glucan strand through its tunnel. This single b-glucan then can be easily hydrolysed by endo-/exo-glucanase. Atomic force microscopy could verify this glucan helicase functionality CFEM-Proteins show in phylogenetic analysis distinct groups with cluster specific functions. That is in accordance with structural information from the protein structure of Candida albicans Csa2 a Pga7-like protein. This group of Pga7-like proteins is involved in heme-iron acquisition from haemoglobin and the transport inside the cell. A time dependent heme binding mechanism could be observed for Pga-like proteins with a two-step binding mode and a structural reorganisation within the binding pocket. Ccw14-like CFEM-proteins lack the heme-binding functionality. This group plays a role in cross-linking the inner cell wall compartment. The interaction of Ccw14 from Saccharomyces cerevisiae with Bgl2, an endo-glucanase could solve the mystery about this group.