Characterization of Chitinases and β-1,3-glucanases in Grapefruit Flavedo during Fruit Development

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Chitinases (EC 3.2.1.14) and β-1,3-endoglucanase (EC 3.2.1.39) are enzymes that are believed to be components of a plant's natural defense against fungal pathogens. We are interested in the potential role of these enzymes in citrus decay resistance. In our preliminary work, we have determined the activities of chitinases and β-1,3-endoglucanases in the flavedo of grapefruit (Citrus paradisi cv. Marsh) 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, β-1,3-endoglucanase activity is lowest in young fruit and increases during development. Western blots of crude flavedo extracts following SDS-PAGE were probed with antibodies raised against purified citrus chitinase and glucanase, and results revealed that changes in the activities of chitinase and β-1,3-endoglucanase were reflected in the amount of chitinase and glucanase protein present in the extracts. Partial purification of flavedo chitinases and glucanases revealed that acidic and basic forms of both enzymes were present in the extracts, although the basic forms were predominant.