Pawpaw [Asimina triloba (L.) Dunal] Fruit Ripening. II. Activity of Selected Cell-wall Degrading Enzymes

Pawpaw fruit were harvested at the advent of the ripening process and were ripened at room temperature. Based on fruit fi rmness and respiration and ethylene production rates at harvest and during ripening, fruit were classifi ed into one of four categories: preripening (no to very slight loss of fi rmness; preclimacteric), early ripening (some softening; increasing rates of ethylene and CO2 production), mid-ripening (soft; at or just past climacteric), and late ripening (very soft; postclimacteric). The activities of the cell-wall degrading enzymes polygalacturonase (PG), endo(1→4)ß-D-glucanase (EGase), and endo-ß-1,4-mannanase (MAN) were low in the preripening and early ripening stages, increased dramatically by mid-ripening coincident with the respiratory and ethylene climacterics, and decreased at late ripening. However, pectin methylesterase (PME) activity per milligram protein was highest at the green stage when the fruit fi rmness was high and decreased as ripening progressed. Tissue prints indicated both EGase and MAN increased as ripening proceeded. The EGase activity was evident near the seeds and the surface of the fruit at preripening and eventually spread throughout, while MAN activity was evident near the fruit surface at preripening and was progressively expressed throughout the fl esh as fruit ripened. The greatest decline in fruit fi rmness occurred between preand early ripening, before the peak activities of PG, EGase, and MAN, although MAN exhibited the greatest relative increase of the three enzymes in this period. The data suggest that PME may act fi rst to demethylate polygalacturonate and may be followed by the action of the other enzymes resulting in cell wall disassembly and fruit softening in pawpaw. Pawpaw fruit soften rapidly after harvest (Archbold and Pomper, 2003; McGrath and Karahadian, 1994), and this hinders the development of effi cient and rapid handling and marketing strategies for the fruit. Softening is an important part of the ripening process, and it is well documented that changes in cell walls accompany fruit softening (Brummell and Harpster, 2001; Seymour and Gross, 1996). Plant cell walls are highly complex structures consisting of many types of complex polysaccharides and numerous proteins (Carpita and Gibeaut, 1993). Pectins, hemicelluloses, and cellulose comprise the major groups of polysaccharides in plant cell walls (Seymour and Gross, 1996), with more than 50% as pectin (Brummell and Harpster, 2001). Cell wall metabolism is tightly controlled through regulation of both gene expression and the cell wall microenviroment (Seymour and Gross, 1996). Due to the structural complexity of the primary cell wall, a coordinated effort of a variety of enzymes affecting both covalent and noncovalent bonds is likely involved in cell wall disassembly and fruit softening (Giovannoni et al., 1992). Other species within the Annonaceae family that includes pawpaw also soften rapidly once ripening begins (Brown et al., Received for publication 26 May 2004. Accepted for publication 14 Jan. 2005. 1Former: Graduate Student, Dept. of Horticulture, Univ. of Kentucky, Lexington,