Although aegerolysin proteins are commonly found in fungi, they can also be found in other organisms. Fungal aegerolysins typically form two-component cytolitic membrane complexes, where component A (aegerolysin) binds to certain membrane lipids or lipid mixtures. The partner proteins of aegerolysins (components B) are proteins with a membrane attack complex perforin (MACPF) domain, that bind to aegerolysins and form a pore in the cell membrane, leading to the cell lysis. The two-component cytolytic complex mucilysin from the mushroom Mucidula mucida was investigated in this MSc thesis. This cytolytic complex consists of mucilysin A (MucA) and mucilysin B (MucB). Mucilysin A belongs to the aegerolysin family, while MucB belongs to proteins with a MACPF domain. Mucilysin A differs from other egerolysins by its higher isoelectric point and by the lack of glutamate in the amino acid sequence (E69), which is important for binding to membranes rich in sphingomyelin (SM) and cholesterol (Chol). Mucilysin B is larger and has a higher isoelectric point and higher molecular weight than other known fungal proteins with a MACPF domain. The genes for mucilysin A and mucilysin B proteins were inserted into plasmid vectors, which were then introduced into E. coli DH5α. Obtained plasmids were then introduced into E. coli BL21(DE3), from which recombinant proteins were isolated. Since we were unable to obtain mucilysin A, further biochemical characterisation tests were performed with already known mushroom aegerolysins. These protein pairs were used in the hemolysis test with bovine erythrocytes at different pH values, with the addition of oxidant and reductant and with the addition of salt. We did not observe hemolysis with any of the tested protein pairs.
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