Purification and characterization of a novel anti-fungal protein from Gastrodia elata

An anti-fungal protein GAFP-1 (Gastrodia anti-fungal protein, also called gastrodianin) was purified from Gastrodia elata B1. f. flavida S. Chow (Orchidaceae), a parasitic plant on the fungus Armillaria mellea. It can inhibit the hyphal growth of some phytopathogenic fungi such as Valsa ambiens, Rhizoctonia solani, Gibberella zeae, Ganoderma lucidum and Botrytis cinerea in vitro. GAFP-1 is a monomer with a molecular mass of 10 kDa and a pI of 8.45. The optimum pH for its inhibitory activity is 5.0 ∼ 6.0. GAFP-1 is insensitive to high temperatures. It can preserve 75% inhibitory activity after 30 min at 60°C. Amino acid composition analysis revealed that GAFP-1 is rich in Asp (22.1%), Gly (10.0 %) and Leu (9.4 %), and does not contain any Pro. The amino acid sequence of the N-terminal was determined and found to share high homology with those of other lectins from orchids such as Listera ovata and Epipactis helleborine. GAFP-1 could not agglutinate trypsin-treated rabbit erythrocytes. It could bind to chitin, immobilized mannose and N-acetylglucosamine in 50mM sodium acetate buffer (pH 5.0) with 2 M ammonium sulfate. These data suggest that GAFP-1 could be a lectin-like protein with strong inhibitory activity against certain fungal pathogens.