Submitted to: Phytopathology
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: August 14, 1996
Publication Date: N/A
Interpretive Summary: The loss of apple fruit in storage is substantial due to decay caused by postharvest pathogens. Although fungicides can effectively control many of these pathogens, public concerns about health and environmental impact may limit their future application. To reduce storage losses and provide a continued source of plentiful food for the consumer, a better understanding gof the mechanism by which postharvest fungal pathogens decay fruit is important in order to develop alternate methods of control. The results of this research indicate that there is only one isozyme of polygalacturonase, a cell wall degrading enzyme produced by Penicillium expansum, the most important postharvest apple pathogen, present in decayed apple fruit. We have purified and characterized this enzyme and are now seeking naturally occurring proteins in the apple fruit which may inhibit the activity of this enzyme. If we can then genetically overexpress this inhibitory protein, it could be used as an alternate method to control postharvest diseases. Apple packing and storage facilities may then be able to reduce the amount of fungicides used to maintain fruit quality in storage.
A polygalacturonase (PG) was purified from rotted cortical tissue of 'Golden Delicious' apple fruit inoculated with Penicillium expansum. The purified protein had a pI of 8.10 and a molecular mass of 34 kDa. The PG was heat labile and most active at pH 5.5. There were no detectable O- or N-linked glycans associated with the PG polypeptide. The purified enzyme macerated apple tissue in situ and in vitro. PG activity was not affected by purified apple polygalacturonase-inhibiting protein (PGIP). Based on the amino acid sequences determined, three degenerate oligonucleotides were synthesized and used as primers to amplify the flanking regions on the fungal genome by polymerase chain reaction (PCR). Amino acid sequences predicted from the cloned PCR products matched perfectly with the determined amino acid sequences, indicating the fungal origin of purified PG.