Plant defensins are cysteine rich cationic
Plant defensins are cysteine-rich cationic peptides of 5–8 kDa whose mature domain comprises 45 to 54 amino ACSF residues, with an isoelectric point around 9. The global fold of plant defensins consists of a cysteine-stabilized motif (CSα/β) made up of an α-helix and a triple-stranded β-sheet stabilized by four disulfide bridges (C1-C8/C2-C5/C3-C6/C4-C7) (Lay et al., 2003). In addition to this motif, Yount et al. (2004) reported the importance of an additional conserved motif, named γ-core, for the antimicrobial activity of the peptide.
The mode of action of several plant defensins has been extensively studied and different mechanistic steps have been identified but are yet not completely understood (Thevissen et al., 2004, Thomma et al., 2002). In many cases, it involves the recognition and binding to a cell wall (e.g., glucosylceramide) or membrane (e.g., phosphatidic acid) receptor. Then, plant defensins are either internalized into the fungal cell or stay at the surface of the membrane and induce several mechanisms: activation of enzyme pathways, modification of ion fluxes, inhibition of conidial germination or induction of apoptosis. In contrast to mammalian and insect defensins, the insertion of plant defensins in the fungal membrane via pore-formation is uncommon and the mechanism of translocation of the peptides has not yet been identified. Plant defensins are generally non-cytotoxic to mammalian and plant cells (Thevissen et al., 2004). However, some plant defensins have been reported to display Ca2+ channel inhibition, anticancer activity and mitogenic activity towards mammalian cell lines (Guzman-Rodriguez et al., 2015). The toxicity of defensins and cAMPs in general has been attributed to the presence of disulfide bonds, their high hydrophobicity and amphipathicity (Hollmann et al., 2016, Jenssen et al., 2006). Consequently, the design of linear analogues of AMPs and defensins showing antimicrobial properties has gained strong interest. For example, a linear variant of human β-defensin 3 has been shown to partially retain its antimicrobial activity, without any toxicity towards human erythrocytes and conjunctiva epithelial cells (Liu et al., 2008).
Fungal spoilage is a major issue in cereal-based foods and in the wider food industry. In addition to economic losses due to unpleasant flavours and appearance, mould has the potential to cause health issues due to the production of mycotoxins (Pitt and Hocking, 2012). Among the different cereal-based products, chilled dough is recognized as a convenient solution to the consumer's desire for oven-fresh baked products. Despite the chilled conditions, bacterial and fungal growth can appear during the storage of this dough. In order to prevent spoilage, a range of preservatives are used, but the overuse of these artificial preservatives has a negative image with the consumer. Alternative solutions to such preservatives, including natural antimicrobial peptides such as nisin, are already in use but their application is still challenging, mainly due to their narrow spectrum of activity (Dielbandhoesing et al., 1998). In contrast, due to their natural role in plant protection, defensins present promising properties, such as inhibition of a broad range of microorganisms and this can be exploited in food applications.
The defensin Cp-thionin II has been extracted from cowpea seeds and displayed antimicrobial activity against Gram-positive and Gram-negative bacteria (Franco et al., 2006). The objective of this study was to characterize the antifungal activity of a linear analogue of Cp-thionin II, KT43C (see Table 1). The application of this synthetic peptide for the prevention of fungal growth in dough was also studied. In addition, the effect of the peptide on red blood cell integrity was examined to determine its toxicity.
Material and methods
Discussion Previous studies have highlighted the antifungal (Carvalho et al., 2001) and antibacterial (Franco et al., 2006) activity of antimicrobial peptides isolated from cowpea seeds, such as the defensin, Cp-thionin II as described by Franco et al. (2006). In this study, a synthetic linear analogue of Cp-thionin II, namely KT43C, exhibited antifungal activity against F. culmorum, P. expansum and A. niger (Fig. 1); while KT43C has been previously shown to inhibit Gram-positive bacteria (Kraszewska et al., 2016), to our knowledge, the antifungal activity of cowpea antimicrobial peptides or derivatives has never been described for the fungi studied here. A synthetic linear analogue of Cp-thionin II, namely KT43C, exhibited antifungal activity against F. culmorum, P. expansum and A. niger (Fig. 1).