Selective mode of action of cucurbitacin in the inhibition of laccase formation inBotrytis cinerea

Polyclonal antibodies were raised against LAC1, one of two isoelectric forms of laccase, and the antibodies, which recognized both of the laccases, LAC1 and LAC2, were then used to demonstrate that both forms have the same molecular range, 72 000M_r. A procedure was developed for labelling laccase at the stage of its secretion into the culture medium, using radioactive leucine. Immuno-precipitation of the [ ¹⁴C]-labelled laccase showed that synthesis of the protein of laccase under inducing conditions was inhibited by cucurbitacins. The degree of inhibition of synthesis of the enzyme protein corresponded to the amount of reduction of laccase activity in the culture medium. Tritiated 2,16,25-cucurbitacin D triacetate was taken up rapidly by young mycelium and appears to be effective as an inhibitor of formation of laccase at micromolar concentrations at the site of action.