Purification and characterization of a pectin lyase from Pythium splendens infected cucumber fruits

A pectin lyase (PL, EC 4.2.2.10) was produced by Pythium splendens Braun in infected cucumber fruits. The enzyme was purified to near homogeneity by ammonium sulfate fractionation, CM - Sepharose column chroma- tography, and preparative isoelectric focusing. The enzyme had a molecular mass of 23 kDa as determined by SDS- PAGE. The pI of the PL was 8.0, which was also the optimum pH for the enzyme activity. The enzyme was stable at 4n50∞C, but beyond 50∞C, its activity decreased rapidly. The pectin lyase degraded high methyl-esterified pectin by trans-elimination in an endo-manner, and was stimulated by some divalent cations including Ca 2+ , Mg 2+ , and Sr 2+ . Addition of Zn 2+ inhibited the enzyme activity by 36%. The purified PL caused rapid maceration of potato and cucumber discs. This is the first report of a PL produced by Pythium splendens, a plant pathogenic fungus which causes seedling damping-off and fruit rot in cucumber.