pH- and concentration-dependent supramolecular assembly of a fungal defensin plectasin variant into helical non-amyloid fibrils

Here the authors report the cryo-EM structure of a triple-mutant of the anti-microbial peptide plectasin, PPI42, assembling in a pH- and concentration dependent manner into helical non-amyloid fibrils. The fibrils formation is reversible, and follows a sigmoidal kinetics. The fibrils adopt a right-handed helical superstructure composed by two protofilaments, stabilized by an outer hydrophobic ring and an inner hydrophobic centre. These findings reveal that alpha/beta proteins can natively assemble into fibrils. Self-assembly and fibril formation play important roles in protein behaviour. Amyloid fibril formation is well-studied due to its role in neurodegenerative diseases and characterized by refolding of the protein into predominantly beta-sheet form. However, much less is known about the assembly of proteins into other types of supramolecular structures. Using cryo-electron microscopy at a resolution of 1.97 angstrom, we show that a triple-mutant of the anti-microbial peptide plectasin, PPI42, assembles into helical non-amyloid fibrils. The in vitro anti-microbial activity was determined and shown to be enhanced compared to the wildtype. Plectasin contains a cysteine-stabilised alpha-helix-beta-sheet structure, which remains intact upon fibril formation. Two protofilaments form a right-handed protein fibril. The fibril formation is reversible and follows sigmoidal kinetics with a pH- and concentration dependent equilibrium between soluble monomer and protein fibril. This high-resolution structure reveals that alpha/beta proteins can natively assemble into fibrils.