AN ANTIMICROBIAL PEPTIDE FROM THE AUSTRALIAN NATIVE HARDENBERGIA-VIOLACEA PROVIDES THE FIRST FUNCTIONALLY CHARACTERIZED MEMBER OF A SUBFAMILY OF PLANT DEFENSINS

An antimicrobial peptide (HvAMP1) was isolated from seeds of the Austr alian native legume Hardenbergia violacea (Schneev.) Stearn. The pepti de 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 inhibit ed the growth of several plant pathogenic fungi at concentrations as l ow as 1 mu M in vitro and produced distinct hyphal distortion and incr eased branching. This antimicrobial activity was greatly diminished in the presence of 1 mM CaCl2 and 50 mM KCl. The purified peptide at 40 mu M did not inhibit three different alpha-amylase enzymes. A eukaryot ic cell-free translation system showed inhibition approaching 50% in t he presence of similar to 100 mu M of HvAMP1. The viability of plant a nd mammalian cells cultured in vitro was not adversely affected by con centrations of HvAMP1 as high as 40 mM. The amino acid sequence of HvA MP1 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 fr om defensins of Vigna unguiculata and Pisum sativum, respectively. Oth er plant defensin sequences showed less than 33% amino acid identity t o the peptide. Therefore, HvAMP1 and the putative plant defensins of c owpea 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 charact erised. The antimicrobial activity of HvAMP1 suggests a defensive role for this subfamily of peptides.