Valerie A. Paynter

Characterization of the Pectic Enzymes from Monilinia fructicola1)

Summary Monilinia fructicola was grown in a chemically defined medium under conditions to give maximum pectic enzyme production. Endo-polygalacturonase (PG), exo-PG, endo-polymethylgalacturonase (PMG), endo-polymethylgalacturonate lyase (PMGL) and pectin methylesterase (PME) activities were detected in the culture filtrates. The PG exhibited a p H optimum at 5.2 and was stable at acidic p H. The PMG exhibited p H optima at 4.5 to 5.0 and 7.5. A heat stability phenomenon was associated with the PG of the two day old culture filtrate.

Partial Purification and Characterization of an Endo-Polygalacturonase from Monilinia fructicola and its Implication in the Brown Rot Disease of Peaches)

Summary A polygalacturonase (PG) from cultural media of Monilinia fructicola was purified 50-fold with 30% yield by ultrafiltration followed by successive chromatography on Sephadex G-75, Ecteola cellulose, and Biogel P-150 columns. The enzyme moved as a single band on disc gel electrophoresis and exhibited a single symmetrical peak on ultracentrifugation. Optimal PG 4 ) activity was at pH 5.2 and 50 ˚C with good enzyme stability between pH 4 and 6. The active form of the enzyme appeared to be a tetramer of molecular weight near 80,000. The enzyme was stable at acid pH but dissociated at pH > 9 to form an inactive monomer with a molecular weight of 20,000. Disc gel electrophoretic patterns supported this hypothesis. Kinetic studies indicated a V max of 2,700 µmoles/min/mg protein and a km of 1 x 10- 6 M with sodium polypectate as substrate. Although viscosity and uronic acid dehydrogenase measurements indicated the endo nature of this PG, hydrolysis of pectate yielded galacturonic acid as the major end product. Anthocyanins extracted from grapes, peaches, and cranberries caused varied inhibition while various polyphenol compounds and benlate, a fungicide, did not inhibit this PG at all. The properties of this enzyme in relation to host-pathogen interaction were discussed.