Purification, characterization and gene cloning of the killer toxin produced by the marine-derived yeast Williopsis saturnus WC91-2

Abstract

As the killer toxin produced by Williopsis saturnus WC91-2 could kill many sensitive yeast strains, including the pathogenic ones, the extracellular killer toxin in the supernatant of cell culture of the marine yeast strain was purified and characterized. The molecular mass of the purified killer toxin was estimated to be 11.0 kDa according to the data from SDS-PAGE. The purified killer toxin had killing activity, but could not hydrolyze laminarin. The optimal conditions for action of the purified killer toxin against the pathogenic yeast Metschnikowia bicuspidate WCY were the assay medium with 10% NaCl, pH 3-3.5 and temperature 16 °C. The gene encoding the killer toxin from the marine killer yeast WC91-2 was cloned and the ORF of the gene was 378 bp. The deduced protein from the cloned gene encoding the killer toxin had 125 amino acids with calculated molecular weight of 11.6 kDa. It was also found that the N-terminal amino acid sequence of the purified killer toxin had the same corresponding sequence deduced from the cloned killer toxin gene in this marine yeast, confirming that the purified killer toxin was indeed encoded by the cloned gene.