Changes in Polygalacturonase Isoenzymes during the ‘Ripening’ of Normal and Mutant Tomato Fruit

Abstract

Polygalacturonase activity is not detectable in mature green tomato fruits but appears as fruits begin to change color and continues to increase during the ripening period. There is a sequential appearance of 2 isoenzymes, polygalacturonase 1 and 2, during ripening. These isoenzymes were purified and their properties compared. Polygalacturonase 1 has a MW of 100,000, is 50% inactivated at 78.degree. C and has a density of 1.343 g cm-3 in cesium chloride. Polygalacturonase 2 has a MW of 42000, is 50% inactivated at 57.degree. C and has a density of 1.300 g cm-3 in cesium chloride. Fruits from isogenic lines homozygous for the ''Neverripe'' (Nr) mutation do not ripen normally and contain reduced amounts of polygalacturonase. Only polygalacturonase 1 is produced in Nr fruit. Tomatoes from isogenic lines homozygous for the ripening inhibitor (rin) mutation do not ripen normally and produce very little detectable polygalacturonase. Although polygalacturonases 1 and 2 have different properties they both give rise to a single polypeptide on electrophoresis in polyacrylamide gels in the presence of sodium dodecylsulfate (MW = 46000). A comparison of the major fragments produced by limited proteolysis of polygalacturonase 1 and 2 with chymotrypsin suggests that the polypeptides from the 2 isoenzymes are similar. The same conclusion was reached from a comparison of polygalacturonase 1 and 2 by radioimmunoassay, using antibody prepared against polygalacturonase 2 and 125I-labeled polygalacturonase 2. The results from radioimmunoassay of extracts from green and ripening fruits suggest that the increase in polygalacturonase activity during ripening is due to net synthesis of protein.