Submitted to: Phytopathology
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: November 21, 2003
Publication Date: April 1, 2004
Citation: Fish, W.W., Davis, A.R. 2004. The purification, physical/chemical characterization, and cDNA sequence of cantaloupe fruit polygalcaturonase-inhibiting protein. Phytopathology. 94(4):337-344.
Interpretive Summary: A conservative estimate of cantaloupe losses in the U.S. domestic market as a result of postharvest decay exceeds $19 million annually. Information from other plant systems suggests that there exists in cucurbits natural inhibitors of fungal plant cell degrading enzymes. The aim of this research was to isolate and characterize one of these natural inhibitors from cantaloupe in order to understand the mechanisms by which it interacts with certain degradative enzymes from important postharvest fungal pathogens. This manuscript describes the isolation, physical/chemical characterization, and cDNA sequence of a polygalacturonase-inhibiting protein from immature cantaloupe fruit. Knowledge about this protein's properties and the mechanism by which it interacts with- and inhibits certain pathogenic fungal enzymes may be utilized to provide natural, effective, pesticide-free control of postharvest decay of netted melons.
Polygalacturonase-inhibiting proteins (PGIPs) are believed to aid in plant defense against fungal pathogens by inhibiting polygalacturonase(s) (PGs) secreted by the invading fungus. In an effort to better understand this type of plant-pathogen interaction in cucurbits, we have isolated a cantaloupe PGIP (CmPGIP) to >90% purity from 5-15 day postanthesis cantaloupe fruit. CmPGIP inhibited crude extracts of PG from two of four cantaloupe fungal pathogens tested. Results from assays for PG activity that utilized rate of substrate viscosity reduction or rate of reducing group formation were consistent with CmPGIP inhibition of endo-PG activity. The Mr of CmPGIP by sedimentation equilibrium or MALDI-TOF MS was 38,500. The pI of CmPGIP was ~8.2, and its absorptivity at 280nm was 0.93 ml/mg. The cd spectrum of native CmPGIP exhibited strong negative ellipticity in the near uv and possessed a far uv spectrum indicative of b-sheet periodic structure. N-terminal amino acid sequence determination of the whole protein and an isolated cyanogen bromide peptide were used to construct oligonucleotide primers for PCR sequencing. The sequenced open reading frame predicts a mature protein of 307 amino acids with up to 68% identity to other PGIP molecules. Northern blot analysis revealed differential expression during fruit development.