Characterization of Hydroxyproline-Containing Hairpin-Like Antimicrobial Peptide EcAMP1-Hyp from Barnyard Grass (Echinochloa crusgalli L.) Seeds: Structural Identification and Comparative Analysis of Antifungal Activity

Herein, we describe a modified form of the antimicrobial hairpin-like peptide EcAMP1, isolated from barnyard grass (E. crusgalli) seeds, which is structurally characterized by a combination of high-pressure liquid chromatography, mass spectrometry, and automated Edman sequencing. This derivate has a single amino acid substitution (Pro19Hyp) in the second α-helical region of the molecule, which is critical for the formation of the hydrophobic core and the secondary structure elements. Comparing the antifungal activity of these two peptides, we found that the modified EcAMP1-Hyp had a significantly weaker activity towards the most-sensitive plant pathogenic fungus Fusarium solani. Molecular dynamics simulations and in vitro binding to the commercial polysaccharides allowed us to conclude that the Pro-19 residue is important for binding to carbohydrates located in the spore cell wall and it chiefly exhibits a fungistatic action representing the hyphal growth inhibition. These data are novel and significant for understanding a role of α-hairpinins in plant immunity.