Summary A new family of antimicrobial peptides has been discovered in Macadamia integrifolia. The first member of this new family to be purified from nut kernels was a peptide of 45 aa residues, termed MiAMP2c. This peptide inhibited various plant pathogenic fungi in vitro. cDNA clones corresponding to MiAMP2c encoded a 666 aa precursor protein homologous to vicilin 7S globulin proteins. The deduced precursor protein sequence contained a putative hydrophobic N‐terminal signal sequence (28 aa), an extremely hydrophilic N‐proximal region (212 aa), and a C‐terminal region of 426 aa which is represented in all vicilins. The hydrophilic portion of the deduced protein contained the sequence for MiAMP2c as well as three additional segments having the same cysteine spacing pattern as MiAMP2c. Each member of the MiAMP2 family (i.e. MiAMP2a, b, c and d) consisted of approximately 50 amino acids and contained a C‐X‐X‐X‐C‐(10–12)X‐C‐X‐X‐X‐C motif. Sub‐ sequent isolations from seed exudates led to the purification of the predicted family members MiAMP2b and 2d, both of which also exhibited antimicrobial activity in vitro. These results suggest that some vicilins play a role in defence during seed germination.

An antimicrobial peptide with no significant amino acid sequence similarity to previously described peptides has been isolated from the nut kernels of Macadamia integrifolia. The peptide, termed MiAMP1, is highly basic with an estimated pI of 10.1, a mass of 8.1 kDa and contains 76 amino acids including 6 cysteine residues. A cDNA clone containing the entire coding region corresponding to the peptide was obtained. The deduced amino acid sequence of the cDNA indicated a 26‐amino‐acid signal peptide at the N‐terminus of the preprotein. Purified MiAMP1 inhibited the growth of a variety of fungal, oomycete and gram‐positive bacterial phytopathogens in vitro. Some pathogens exhibited close to 100% inhibition in less than 1 μM peptide (5 μg/ml). Antimicrobial activity was diminished against most, but not all, microbes in the presence of calcium and potassium chloride salts (1 mM and 50 mM, respectively). MiAMPl was active against bakers yeast, was inactive against Escherichia coli and was non‐toxic to plant and mammalian cells. Analysis of genomic DNA indicated that MiAMP1 was encoded on a single copy gene containing no introns. The MiAMP1 gene may prove useful in genetic manipulations to increase disease resistance in transgenic plants.

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An antimicrobial peptide (HvAMP1) was isolated from seeds of the Australian native legume Hardenbergia violacea (Schneev.) Stearn. The peptide is 47 amino acid residues in length, contains 8 cysteines, and has a molecular weight of 5392 and a predicted pI of 10.41. HvAMP1 inhibited the growth of several plant pathogenic fungi at concentrations as low as 1 µM in vitro and produced distinct hyphal distortion and increased branching. This antimicrobial activity was greatly diminished in the presence of 1 mM CaCl2 and 50 mM KCl. The purified peptide at 40 µM did not inhibit three different a-amylase enzymes. Aeukaryotic cell-free translation system showed inhibition approaching 50% in the presence of ~100 µM of HvAMP1. The viability of plant and mammalian cells cultured in vitro was not adversely affected by concentrations of HvAMP1 as high as 40 mM. The amino acid sequence of HvAMP1 contained the consensus amino acids that define the plant defensin family of peptides. The HvAMP1 amino acid sequence showed 87% and 57% identity with the amino acid sequences deduced from cDNA sequences from defensins of Vigna unguiculata and Pisum sativum, respectively. Other plant defensin sequences showed less than 33% amino acid identity to the peptide. Therefore, HvAMP1 and the putative plant defensins of cowpea and pea define a distinct sequence subfamily of plant defensins which is at present limited to members of the Fabaceae. HvAMP1 is the first member of this subfamily to be purified and functionally characterised. The antimicrobial activity of HvAMP1 suggests a defensive role for this subfamily of peptides.

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