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Class I Hydrophobin Vmh2 Adopts Atypical Mechanisms to Self-Assemble into Functional Amyloid Fibrils

Gravagnuolo, Alfredo Maria, Longobardi, Sara, Luchini, Alessandra, Appavou, Marie-Sousai, De Stefano, Luca, Notomista, Eugenio, Paduano, Luigi, Giardina, Paola
Biomacromolecules 2016 v.17 no.3 pp. 954-964
Pleurotus ostreatus, amyloid, aqueous solutions, calcium, circular dichroism spectroscopy, fluorescence, fungi, hydrophobins, ions, light scattering, models, pH, solubilization, temperature, transmission electron microscopy
Hydrophobins are fungal proteins whose functions are mainly based on their capability to self-assemble into amphiphilic films at hydrophobic–hydrophilic interfaces (HHI). It is widely accepted that class I hydrophobins form amyloid-like structures, named rodlets, which are hundreds of nanometers long, packed into ordered lateral assemblies and do not exhibit an overall helical structure. We studied the self-assembly of the Class I hydrophobin Vmh2 from Pleurotus ostreatus in aqueous solutions by dynamic light scattering (DLS), thioflavin T (ThT), fluorescence assay, circular dichroism (CD), cryogenic trasmission electron microscopy (cryo-TEM), and TEM. Vmh2 does not form fibrillar aggregates at HHI. It exhibits spherical and fibrillar assemblies whose ratio depends on the protein concentration when freshly solubilized at pH ≥ 7. Moreover, it spontaneously self-assembles into isolated, micrometer long, and twisted amyloid fibrils, observed for the first time in fungal hydrophobins. This process is promoted by acidic pH, temperature, and Ca²⁺ ions. A model of self-assembly into amyloid-like structures has been proposed.