Selected Wheat Seed Defense Proteins Exhibit Competitive Binding to Model Microbial Lipid Interfaces

Puroindolines (Pins) and purothionins (Pths) are basic, amphiphilic, cysteine-rich wheat proteins that play a role in plant defense against microbial pathogens. This study examined the co-adsorption and sequential addition of Pins (Pin-a, Pin-b, and a mutant form of Pin-b with Trp-44 to Arg-44 substitution) and beta-purothionin (beta-Pth) model anionic lipid layers using a combination of surface pressure measurements, external reflection FTIR spectroscopy, and neutron reflectometry. Results highlighted differences in the protein binding mechanisms and in the competitive binding and penetration of lipid layers between respective Pins and beta-Pth. Pin-a formed a blanket-like layer of protein below the lipid surface that resulted in the reduction or inhibition of beta-Pth penetration of the lipid layer. Wild-type Pin-b participated in co-operative binding with beta-Pth, whereas the mutant Pin-b did not bind to the lipid layer in the presence of beta-Pth. The results provide further insight into the role of hydrophobic and cationic amino acid residues in antimicrobial activity.