Snakin-1, a Peptide from Potato That Is Active Against Plant Pathogens

Abstract

A new type of antimicrobial peptide, snakin-1 (SN1), has been isolated from potato tubers and found to be active, at concentrations <10 uM, against bacterial and fungal pathogens from potato and other plant species. The action of SN1 and potato defensin PTH1 was synergistic against the bacterium Clavibacter michiganensis subsp. sepedonicus and additive against the fungus Botrytis cinérea. Snakin-1 causes aggregation of both gram-positive and gram-negative bacteria. The peptide has 63 amino acid residues (M r 6,922), 12 of which are cysteines, and is unrelated to any previously isolated protein, although it is homologous to amino acid sequences deduced from cloned cDNAs that encode gibberellin-inducible mRNAs and has some sequence motifs in common with kistrin and other hemotoxic snake venoms. A degenerate oligonucleotide probé based on the internal sequence CCEECKC has been used to clone an SN1 cDNA. With the cDNA used as probé, one copy of the StSNl gene per haploid genome has been estimated and expression of the gene has been detected in tubers, stems, axillary buds, and young floral buds. Expression levéis in petáis and carpels from fully developed flowers were much higher than in sepáis and stamens. The expression pattern of gene StSNl suggests that protein SN1 may be a component of constitutive defense barriers, especially those of storage and reproductive plant organs.Plants and animáis are in cióse contact with widely diverse bacteria and fungi, but only in rare cases does this association result in the development of disease, substantially because of the existence of defense systems. Although considerable differences exist among different types of organisms with respect to their defense mechanisms, the most notable of which is the lack of an adaptive immune response in plants, recent evidence indicates that both plants and animáis share some com-

Expression, purification and characterization of the recombinant cysteine-rich antimicrobial peptide snakin-1 in Pichia pastoris

“…6, F. oxysporum was highly susceptible to recombinant SN-1 and spore germination was completely inhibited with 60 μM of SN-1. This result was in accordance with the findings described in previous reports for the potato native SN-1 and chemically synthetic SN-1 [19,24].…”

“…Many AMPs are cysteine-stabilized and these have been classified into several families [14], including cyclotides [15], defensins [16], hevein-like peptides [17], lipid transfer proteins [18], and snakins [11,19]. The snakin family is a novel plant antimicrobial peptide family which shows good similarity with the members of the gibberellic acid stimulated transcript (GAST) and gibberellic acid stimulated in Arabidopsis (GASA) protein families in Arabidopsis [20].…”

“…These cysteine-rich peptides are widely distributed among plant species, including potato, tomato, avocado, petunia, French bean, gerbera, strawberry, maize, soybean, pepper, rice, Arabidopsis, and alfalfa [20][21][22][23]. All snakin peptides are characterized by twelve conserved cysteine residues that are involved in the formation of six disulfide bonds [19]. These disulfide bonds are thought to be important for maintaining their tertiary structure and also responsible for their biological activity [13,24,25].…”

“…These disulfide bonds are thought to be important for maintaining their tertiary structure and also responsible for their biological activity [13,24,25]. In previous studies, snakin peptides from potato (Solanum tuberosum) were found to exert strong antimicrobial activity against phytopathogens [11,19,24,26] and animal pathogens [27]. However, little is known about their mechanism of action against target organisms.…”

“…The amino acid sequence of SN-1 is GSNFCDSKCKLCSKAGLADRCLKYCGICCEECKCVPSGTYGNKHECPCYRDKK NSKGKSKCP [19,22]. Computational modeling indicates that the 3-D structure of the SN-1 peptide is composed of two long α-helices with six disulfide bonds and shows a small degree of structural similarity with thionins [25].…”