Cellulase in Tomato Fruits

196

Ultrastructural changes in the pericarp of tomato (Lycpekon escudentun Mill) fruit were followed during ripening. Ethylene production was monitored by gas chromatography and samples analyzed at successive stages of the ripening process.Changes in the cytoplasm}c uktrastructure were not consistent with the suggestion that ripening is a 'senescence' pbenomenon. A large degree of ultrastructural organization, especially of the mitochondria, chromoplasts, and rough endoplasmic reticulum, was retained by ripe fruit.Striking changes in the structure of the cell wall were noted, begnning with dissolution of the middle lamella and eventual disruption of the primary cell wall. These changes were correlated with appearance of polygalacturonase (EC 3.2.1.15) isoenzymes. Application of purifli tomato polygalacturonase lsoenzymes to mature green fruit tissue duplicated the changes in the cell wall noted duing normal ripening. Possible roles of the polygalacturonase isoenzymes in cell wall disorganizadon are discussed.

“…This has been shown to be associated with an increase in the activities of cell wall degrading enzymes, particularly PG2 (5, 7), although a role has been suggested for cellulase (4,11).…”

mentioning

confidence: 99%

Ultrastructure of Tomato Fruit Ripening and the Role of Polygalacturonase Isoenzymes in Cell Wall Degradation

Crookes

Grierson²

1983

196

“…In method A, 3 g of CMC were added to 250 ml of 0.1 N Na acetate buffer at pH 5.0 containing 1 jtg/ml of merthiolate as a preservative. Merthiolate is known not to inhibit tomato fruit cellulase (7,33). The mixture was blended for 3 min in a Waring Blendor at high speed.…”

Section: Methodsmentioning

confidence: 99%

“…These include hormonally induced growth responses (6,10), abscission of leaves (1,2,14,19), pollen tube growth (16,31), and softening of fruits as they ripen (7,12,33). However, little is known about the nature of cellulase from higher plants despite the probable physiological importance of cellulase in growth and development.…”

mentioning

confidence: 99%

Partial Characterization of C_x Cellulase and Cellobiase from Ripening Tomato Fruits

Pharr¹,

Dickinson²

1973

Self Cite

Cellulolytic enzymes were studied in extracts from the locular contents of ripening fruits of Lycopersicon esculentum var. KC-146. When acting on carboxymethyl cellulose, the enzyme preparations were capable of decreasing the viscosity of the reaction mixture and generating reducing groups, oligosaccharides, and glucose. Cellobiose celiotriose, celiotetrose, and cellopentose also served as substrates for glucose production.

“…Cellulolytic enzymes increase in activity in some fruits during ripening (7,9,10,17), in abscission zones of petioles prior to leaf abscission (1,3,12,13), in rapidly expanding epicotyls after auxin treatment (8,16,19), and appear to be involved in pollen tube elongation (22 15), an increase in cellulase activity has been detected only after auxin treatment (16,19,20). The present work provides evidence that in the expanding pea epicotyl, auxin and ethylene each appear to control the development of a different form of cellulase and each enzyme possesses different requirements for activity and apparently differing subcellular locations and isoenzyme complements.…”

Section: Introductionmentioning

confidence: 99%

mentioning

confidence: 99%

Extraction and Partial Characterization of Cellulases from Expanding Pea Epicotyls

Ferrari

Arnison

1974

Etiolated pea (Pisum sativum) epicotyls synthesize a buffersoluble cellulase (cellulase A) and a salt-soluble cellulase (cellulase B) (EC 3.2.1.4) after treatment with high (0.5%) auxin levels. Only cellulase A increased in activity after treatment with low (0.005 % ) auxin. Cellulase A was released into the supernatant after homogenization of tissue in dilute buffer (buffer-soluble), had a pH optimum at 5.5, was relatively thermostable, and its activity was inhibited by NaCl. Cellulase B was released by 1 M NaCl (salt-soluble) from excised tissue segments or from the insoluble residue remaining after removal of the buffer-soluble form. It had a pH optimum at 7.0, was thermolabile, and required salt for maximum activity. When subjected to polyacrylamide gel electrophoresis, the cellulase fraction released by NaCl from excised segments showed two bands of celiulase activity compared to several for the buffer-soluble fraction. Electrophoretic analysis of the buffer and salt-soluble fractions for marker enzymes indicated the presence of malate dehydrogenase activity in all fractions and glutamate dehydrogenase activity in the buffer-soluble fraction only.Exposure of intact pea epicotyls to 70 ,ml/I of ethylene gas for 3 days did not affect cellulase A activity, but caused a 5-fold increase in cellulase B activity (enzyme released by salt from the buffer-insoluble residue). We concluded that ethylene and auxin generate different forms of cellulase.Cellulolytic enzymes increase in activity in some fruits during ripening (7,9,10,17), in abscission zones of petioles prior to leaf abscission (1,3,12,13), in rapidly expanding epicotyls after auxin treatment (8,16,19), and appear to be involved in pollen tube elongation (22 15), an increase in cellulase activity has been detected only after auxin treatment (16,19,20). The present work provides evidence that in the expanding pea epicotyl, auxin and ethylene each appear to control the development of a different form of cellulase and each enzyme possesses different requirements for activity and apparently differing subcellular locations and isoenzyme complements. MATERIALS AND METHODSAfter 7 to 8 days growth, etiolated pea (Pisum sativum) seedlings were decapitated and the epicotyls were treated with auxin (indoleacetic acid 0.5%, except where indicated) as described previously (8). Two to 4 days later, salt-soluble and buffersoluble cellulases were extracted from apical, swollen tissue segments which were approximately 10 mm in length. Tissue samples (1 g) were homogenized with mortar and pestle in 20 mm sodium phosphate buffer (pH 6.5, 1 g/ 2 ml of medium) containing 0.02% NaN. (as a precaution against microbial contamination). The homogenate was centrifuged (37,000g, 10 min), and the supernatant containing the buffer-soluble cellulase fraction was saved. The pellet (buffer-insoluble residue) was washed twice in homogenization medium (1 ml/g fresh weight of tissue), and after centrifugation, the supernatants were combined. (In the experiment presented in Table I, t...