Purification and characterisation of a prominent polygalacturonase isozymeproduced by Phomopsis cucurbitae in decayed muskmelon fruit

Phomopsis cucurbitae is a latent infection fungus which causes muskmelon fr uit decay only after harvest. It can cause severe losses during fruit stora ge and marketing in the U.S.A., Japan and some Central American countries. Previous studies showed that P. cucurbitae produced polygalacturonase (PG) in both culture and muskmelon fruit tissue. In this study, a prominent PG i sozyme (PGI) produced by P. cucurbitae in decayed fruit was purified to hom ogeneity by a procedure of extraction ultrafiltration, preparative isoelect ric focusing, anion exchange, and gel filtration chromatography. The isozym e had a primary endo-activity, a relative molecular weight of 54 kD accordi ng to SDS-PAGE, and a pi of 4.2 based on IEF-PAGE. The isozyme was active i n a low pH range from 2 to 6, and was optimum at pH 5. Optimum temperature for the activity of PGI was 40-45 degrees C. The K-m and V-max on polygalac turonic acid were 44.7 mu g ml(-1) and 0.14 U mu g(-1) enzyme, respectively . The purified PGI effectively macerated mature fruit tissue, suggesting th at the isozyme may be involved in the pathogenesis of Phomopsis fruit rot o f muskmelon.