Purification of pectate lyase produced by Colletotrichum gloeosporioides and its inhibition by epicatechin: a possible factor involved in the resistance of unripe avocado fruits to anthracnose.

Pectate lyase (PL) from Colletotrichum gloeosporioides was purified to apparent homogeneity by hydrophobic interaction chromatography followed by isoelectric focusing. The purified preparation showed one band corresponding to 40 kD on sodium dodecyl sulfate-polyacrylamide gels. The isoelectric point of the enzyme was 7.9, and the optimum pH for activity was 8.9. The purified PL efficiently macerated unripe avocado fruit wedges. In vitro translation of mRNA from an induced fungal culture revealed a 36-kD precursor polypeptide, which was precipitated with PL antibodies. The antibodies inhibited enzymatic activity and maceration ability on avocado wedges. Epicatechin, a flavan 3-ol present in the peel of unripe avocado fruit, had a Ki of 3.4 microM for inhibition of PL activity in vitro. At 20 micrograms/ml (68 microM), epicatechin reduced the enzyme's macerating ability by 64%. Since the flavan is present in unripe fruit at much higher concentrations (about 350 micrograms/g fresh weight) than the inhibitory concentrations, epicatechin may be involved in the resistance of unripe avocado fruits by inhibiting the PL activity of C. gloeosporioides.