Properties of Rhizopus stolonifer Polygalacturonase, an Elicitor of Casbene Synthetase Activity in Castor Bean (Ricinus communis L.) Seedlings.
Some properties of the polygalacturonase-elicitor from the filtrates of Rhizopus stolonifer cultures have been examined in an attempt to understand its mode of action as an elicitor of casbene synthetase activity in castor bean seedlings. Both the polygalacturonase activity and the elicitor activity are heat-labile with similar heat-sensitivity profiles. Also, the catalytic activity of the enzyme is lost on treatment with sodium periodate, as had been shown previously for the elicitor activity. The pH optimum of the enzyme activity with polygalacturonic acid as the substrate is 4.9. Exposures of germinating castor bean seedlings to the elicitor for short-term periods of 1 to 10 minutes followed by washing and incubation in sterile, distilled water are partially effective in elicitation in comparison with the continuous exposure of the seedlings over 11 hours to the same amount of the elicitor. The initial rate of reaction catalyzed by the enzyme is about 3 times faster with polygalacturonic acid as a substrate than with partially (50%) methylated polygalacturonic acid (pectin). The K(m) value of the enzyme for polygalacturonic acid is about 4.2 millimolar in terms of monomeric units and about 0.07 millimolar in terms of polymer concentration. Examination of the types of products formed by the action of the enzyme suggests that it is an endo-hydrolase. The amino acid composition of this enzyme is similar to those of other extracellular fungal proteins reported. The carbohydrate moiety of the glycoprotein polygalacturonase-elicitor is composed of 92% mannose and 8% glucosamine by gas chromatography-mass spectrometry analysis. The linkage group analysis of the carbohydrate moiety showed that mannosyl residues which are 1,2-linked comprise about 70% of the total glycosyl residues and demonstrated the presence of some 1,3,6- and 1,2,6-linked branching mannosyl residues.
Lee
S C
SC
Division of Biochemistry, Department of Chemistry, University of California, Los Angeles, California 90024.
West
C A
CA