# Towards Understanding the Function of Aegerolysins
## 3 Sequences, Structures, and Structural Models of Aegerolysins  
We have collected published experimental data on several aegerolysins and their analogs. The sequences of these aegerolysins were taken from published papers or, if not available there, from publicly available genomes; to avoid confusion, they were collected in File S1. The sequences of the aegerolysins ranged from 11 to 92% identical. Only three conserved amino acids, all three glycines, were observed in the alignment between them (Figure S1, star in magenta), and their position in the structure of OlyA6 (PDB ID: 6 MYJ) was marked (Figure 2, in magenta). Two of them were located in two different loops on the same side of the molecule; the second loop contains an additional almost conserved glycine (except in **Cry34 Ab1**), followed by a \(\beta\)-sheet containing the third conserved glycine. Due to the smallest side chain, the glycine can increase the local flexibility in the protein structure and the intrinsically destabilize \(\beta\)-sheet [153], possibly helping to accommodate the binding site. In the structure of **OlyA6**, the position of the fragment of the chemical structure of SM (18:1) is marked (Figure 2, in orange and orange arrow) [73]. Genome sequences from multiple strains are available for some of the species. Although we identified only one gene locus encoding the same aegerolysin sequence in the genome of the two laboratory monokaryotic model strains of _P. ostreatus_ PC9 and PC15, the coding sequences of the aegerolysin protein may vary between other strains [52; 53; 54; 55]. For example, the **OlyA6** protein differs in one amino acid (99% identical) from the genomic sequence **PlyA(PriA)**, and **PlyA** in six (95% identical) [52]. Perhaps it is also plausible to expect _P. eryngii_, a grassland litter decomposer consisting of at least five varieties such as _var. eryngii_, _ferulae_, _ealeoselini_, _thapsiae_, and _tingitanus_, which differ significantly with habitat distribution, host, and morphology, to contain analogs of the aegerolysins **PlaA2** or **EryA**. Similar variations in aegerolysin sequences have already been observed in some _Altenaria_ strains [36]. Of the two aegerolysins described in _A. fumigatus_, an invariant **asp-HS-like** sequence was also present in six other strains; variation in one amino acid was observed in **asp-HS** between the strains, but **asp-HS** was also absent in one of the strains (sequence of component B from the same locus, **asp-HSB**, was only partial in the same strain). Variations in the aegerolysin sequences for **NigA1** were not observed in any of the nine other _A. niger_ strains examined. A similar observation was made for **NigA2**, except for a variation of four amino acids in one of the strains. There are some differences in the sequences around the putative intron splice site and at the C terminus among the three _A. terreus_ strains. Four genome sequences of _A. oryzae_ are accessible in public databases; surprisingly, only one aegerolysin with only one amino acid variation is encoded in all four strains. Another strain has an additional gene identical to the strain with three aegerolysin genes. In the last strain, three additional sequences are found, one correspondingly short and two larger, one with a longer C-terminus and the other with a long N-terminus. The sequences of **BlyA** are identical in all four strains of _B. bassiana_. In the sequence of **Ag11**, there is only one amino acid variation among the four _T. atrovidride_ strains. One of the phylogenetically more distant strains, _P. aeruginosa_ PA7 (according to whole genome comparison) revealed that **Rah**U is only 77% identical to the PAO strain. Sequencing and annotation errors cannot be excluded. Figure 2: Phylogenetic tree of aegerolysins with some published information combined with their structures and models.