Solid-phase synthesis, conformational analysis, and biological activity ofAVR(9) elcitor peptides of the fungal tomato pathogen Cladosporium fulvum

The race-specific peptide elicitor AVR9 of the fungal pathogen Cladosporium fulvum specifically induces a hypersensitive response in tomato genotypes carrying the complementary resistance gene Cf-9. The total chemical synthes es of this 28-residue AVR9 peptide containing three disulfide bonds, and of three mutant peptides [R8K]AVR9, [F10A]AVR9 and [F21A]AVR9, have been acco mplished. The syntheses were carried out using a stepwise solid-phase appro ach based on tBoc chemistry. The disulfide bridges were formed by air oxida tion. The correctness of the chemical structure of all folded synthetic pep tides was confirmed by combined NMR and MS analyses. The biological activit y and a number of physicochemical properties of folded synthetic AVR9 are i dentical to those of native fungal 28-residue AVR9. The overall conformatio ns of the folded synthetic mutant peptides were comparable to that of synth etic wild-type AVR9 as demonstrated by NMR spectroscopy. Mutant [R8K]AVR9 s howed a threefold higher, and mutant [F10A]AVR9 a threefold lower necrosis- inducing activity when compared to synthetic wild-type AVR9. However, mutan t [F21A]AVR9 showed hardly any necrosis-inducing activity. Affinity for pol yclonal antibodies raised against native fungal AVR9 is positively correlat ed with the necrosis-inducing activity of the synthetic AVR9 peptides ([R8K ]AVR9 > wild-type AVR9 > [F10A]AVR9 > [F21A]AVR9).