Purification and characterization of an endopolygalacturonase from Verticillium albo-atrum

A polygalacturonase was isolated from the culture filtrate of the fungal pl ant pathogen Verticillium albo-atrum and purified 22-fold to homogeneity as judged by SDS-electrophoresis. The enzyme was a basic protein with a molec ular weight of 37 kDa, an isoelectric point greater than or equal to 8.6 an d containing 1.7% carbohydrate. The enzyme was an endo-polygalacturonase an d hydrolysed a wide range of pectic substrates including polygalacturonic a cid, 93% methylated pectin and pectins in tomato cell walls. The best subst rate was 31% methylated pectin. Relative reaction rates on pectins with dif ferent degrees of methylation could be explained by considering both the nu mber of susceptible bonds and non-specific enzyme-substrate interactions. T he principal products of long-term hydrolysis were di- and mono-galacturona te. Maximum activity was observed at pH 4.6-5.0 and 46 degrees C. However, the enzyme lost activity above 30 degrees C in the absence of substrate. En zyme activity was very sensitive to changes in ionic strength at low salt l evels. It was stable in the pH range 3-11 at 30 degrees C.