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Title: CHARACTERIZATION OF CHITINASES AND B-1,3-GLUCANASES IN GRAPEFRUIT FLAVEDO DURING FRUIT DEVELOPMENT

Author
item McCollum, Thomas
item Doostdar, Hamed
item Mayer, Richard
item McDonald, Roy

Submitted to: Physiologia Plantarum
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 7/2/1996
Publication Date: N/A
Citation: N/A

Interpretive Summary: We are interested in the roles of chitinase and glucanase, two enzymes known to be involved in plant's resistance to fungal and insect pests, in Citrus species. This report presents results of experiments that were designed to determine levels of these enzymes in grapefruit flavedo during development and on partial purification and biochemical characterization of fthe enzymes. We found that levels of the two enzymes vary with tissue age and that their levels are regulated independently. Chitinase activity is highest in young fruit, and decreases as fruit age whereas glucanase activity is lowest in young fruit and increases as fruit age. Results of partial purification and biochemical characterization revealed that multiple forms of the two enzymes are expressed in grapefruit flavedo. Additional research will be required to determine the effects of these particular enzymes on fungal and insect pests. However, our results do suggest that chitinase and glucanase levels may be amenable to genetic manipulation. It is expected that such manipulation may lead to enhanced resistance of citrus to pests.

Technical Abstract: Chitinase (EC 3.2.1.14) and B-1,3-glucanase (EC 3.2.1.39) activities in the flavedo of grapefruit (Citrus paradisi cv. Marsh) were determined at 17 times during the course of fruit development. Chitinase activity is initially high in flavedo, but drops rapidly and is low, although fairly constant throughout the remainder of fruit development. In contrast to chitinase, B-1,3-glucanase activity is lowest in young fruit and increases during development. Results of immunostaining revealed that changes in the activities of chitinase and B-1,3-glucanase were reflected in the amount of chitinase and glucanase protein present in the extracts. Partial purification of flavedo chitinases and glucanases was performed using extracts prepared from immature and mature fruit for the two enzymes, respectively. Acidic and basic forms of both enzymes were present in the extracts; acidic and basic forms of chitinase were present in nearly equal amounts whereas basic glucanases predominated (91% of total activity). Acidic and basic chitinases differed in substrate specificity as well as products of degradation indicating the heterogeneous nature of the enzymes. Both acidic and basic glucanases required the presence of B-1,3 linkages for activity, were active against both soluble and insoluble B-1,3 glucans and generated similar products.