Rice Defensin OsAFP1 is a New Drug Candidate against Human Pathogenic Fungi

Abstract

Fungal infections, such as candidiasis and aspergillosis, are some of the most frequent infections in humans. Although antifungal drugs are available for the treatment of these infections, antifungal agents with new mechanisms of action should be developed because of the increasing incidence of drug-resistant pathogens in recent years. In this study, a basic functional analysis of rice defensin OsAFP1, a novel antifungal drug candidate, was conducted. OsAFP1 exerted fungicidal activity against Candida albicans, the most common pathogenic fungus in humans, at 4 μM concentration, but it did not inhibit the growth of human pathogenic bacteria. In addition, OsAFP1 retained structural stability after heat treatment at 100 °C for 10 min and after serum treatment at 37 °C for 24 h. A propidium iodide (PI) uptake assay and mutational analysis revealed that amino acid residues within the C-terminal γ-core motif of OsAFP1, particularly Leu-39 and Lys-41, play an important role in its antifungal activity. Further, PI uptake and apoptosis assays suggested that OsAFP1 exerts its antifungal activity by inducing apoptosis of target cells. Immunohistochemistry showed that the OsAFP1 target molecule was located in the cell wall. These findings indicate that OsAFP1 may be developed into a potent antifungal drug.

Defensins of Grasses: A Systematic Review

“…The following defensins of rice Oryza sativa L. were studied in detail: OsDEF7 and OsDEF8 by Tantong et al [ 67 ] and Weerawanich et al [ 93 ]; OsDEF7, named OsAFP1, was further studied by Sagehashi et al [ 94 ] and Ochiai et al [ 95 ], while Cal1, identical to OsDEF7, was analyzed by Luo et al [ 96 ].…”

“…Ochiai et al [ 95 ] studied the effect of OsDEF7 on the growth of human pathogenic fungi and showed that the peptide exerted fungicidal activity against Candida albicans at a concentration of 22.2 µg/mL. The recombinant peptide also inhibited the growth of two Saccharomyces cerevisiae strains (MIC = 22.2 µg/mL and 88.8 µg/mL), but failed to inhibit the growth of human pathogenic bacteria, such as Gram-negative bacteria E. coli K-12 and Porphyromonas gingivalis , and Gram-positive bacteria Streptococcus mutans , Staphylococcus aureus , and Propionibacterium acnes at concentrations below 177.7 µg/mL.…”

“…The recombinant peptide also inhibited the growth of two Saccharomyces cerevisiae strains (MIC = 22.2 µg/mL and 88.8 µg/mL), but failed to inhibit the growth of human pathogenic bacteria, such as Gram-negative bacteria E. coli K-12 and Porphyromonas gingivalis , and Gram-positive bacteria Streptococcus mutans , Staphylococcus aureus , and Propionibacterium acnes at concentrations below 177.7 µg/mL. The peptide retained activity after heating at 100 °C for 10 min and serum incubation at 37 °C for 24 h. Analysis of the mode of action of the peptide showed that it binds to phosphatidylinositols of the C. albicans cell membranes and induces apoptosis [ 95 , 97 ].…”

“…Structure–function relationships in rice defensins were studied using OsDEF7 (OsAFP1) as a model [ 94 , 95 ]. Eight short peptides covering the entire sequence of OsDEF7 were synthesized.…”

“…Eight short peptides covering the entire sequence of OsDEF7 were synthesized. The inhibitory activity of the short peptides was assayed against the plant ( P. oryzae ) and human ( C. albicans ) pathogens [ 94 , 95 ]. Peptides 1 (residues 1‒10), 2 (residues 5‒15), 7 (residues 35‒44), and 8 (residues 40‒49) derived from the N-and C-terminal regions of OsDEF7 displayed antimicrobial activity against P. oryzae (IC 50 = 0.41‒1.42 µg/mL).…”