Die Rolle der Phosphorylase im Stoffwechsel der Stärke in den Plastiden

Fekete, M. A. R. de

doi:10.1007/bf00396028

Cited by 46 publications

(7 citation statements)

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“…Multiple forms of phosphorylase have been found in various plant tissues; spinach leaf (3,19,23), bundle sheath cells of maize leaf (25), pea leaves (21,23), banana fruit pulp (21,22), developing and germinating seed of peas (17,18,21), pea roots, broad bean seeds, mature seeds of green beans and mung beans, potato tubers, tobacco leaf and developing seeds of rice (21), banana leaf (11), and in developing maize endosperm (26).…”

Section: Discussionmentioning

confidence: 99%

“…Association of one of the multiple phosphorylase forms with the chloroplast has been shown for only spinach leaf (3,19,23), pea leaf (23), and maize leaf bundle sheath cells (25). The developing and germinating pea seed phosphorylases are the only other isozymes that have been characterized with respect to kinetic studies using various high mol wt glucans (17,18).…”

Section: Discussionmentioning

confidence: 99%

“…Recent studies indicate that multiple forms of phosphorylase do exist in spinach leaf (3,19,23 Preparation of Debranched Amylopectin. Two g amylopectin in 50 ml of 20 mm citrate buffer (pH 5.0) was incubated with 50 ,ug pullulanase (Boehringer Mannheim, 10 IU/mg) at 37 C for 21 h. The debranched amylopectin was then heated at 100 C for 5 min and clarified by centrifugation at 15,000g for 10 min.…”

Section: Introductionmentioning

confidence: 99%

“…Recent studies indicate that multiple forms of phosphorylase do exist in spinach leaf (3,19,23). One of these forms appears to be localized within the chloroplast.…”

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confidence: 99%

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Characterization of the Spinach Leaf Phosphorylases

Preiss¹,

Okita²,

Greenberg³

1980

Plant Physiol.

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The chloroplastic and the cytoplasmic phosphorylases were purified and their kinetic properties characterized. The cytoplasmic enzyme was purified to homogeneity via affinity chromatography on a glycogen-Sepharose column. Subunit molecular weight studies indicated a value of 92,000, whereas a native molecular weight value of 194,000 was obtained by sucrose density gradient centrifugation. The Little is known about the primary steps involved in chloroplastic starch degradation. Recent investigations on starch degradation in isolated spinach chloroplasts have indicated that both maltose and 3-P-glycerate are end products (8,13,20). This suggests that in spinach chloroplasts both amylolytic and phosphorolytic cleavages are occurring. In contrast, negligible amylolytic activity is found in pea chloroplast extracts, and starch degradation appears to occur solely via phosphorolysis (14, 24).Thus far there is little available information on the isolation and characterization of the starch degradative enzymes in spinach leaf. Recent studies indicate that multiple forms of phosphorylase do exist in spinach leaf (3,19,23 Preparation of Debranched Amylopectin. Two g amylopectin in 50 ml of 20 mm citrate buffer (pH 5.0) was incubated with 50 ,ug pullulanase (Boehringer Mannheim, 10 IU/mg) at 37 C for 21 h. The debranched amylopectin was then heated at 100 C for 5 min and clarified by centrifugation at 15,000g for 10 min. The mixture was then diluted with H20 to a final concentration of 10 mg/ml. This preparation contains two maltodextrin fractions; one with a chain length range between 12 to 42 glucosyl residues and the other with chain lengths greater than 49 glucosyl residues (7).Preparation of Partially Hydrolyzed Amylose. Potato amylose type I, 4.5 g, was suspended in 225 ml of 0.1 N HCI and boiled in a water bath for 60 min. The solution was cooled and neutralized with 10 N NaOH to pH 7.0.Sucrose Density Gradient Ultracentrifugation of Phosphorylase. Sucrose density ultracentrifugation was done according to the procedure of Martin and Ames (16). Lactate dehydrogenase (mol wt, 140,000) and pyruvate kinase (mol wt, 237,000) were used as marker enzymes.Preparation of the Glycogen Affinity Resin. An affinity column with glycogen attached to Sepharose 6B was prepared according to the method of Vretblad (27). Epoxy-activated Sepharose 6B (Pharmacia), 15 g, was suspended in 94 ml deionized H20 and washed with 1,500 ml H20 for I h on a coarse glass filter. The gel was washed with 94 ml of 0.1 M NaOH and then transferred to 45 ml of a 0.1 M NaOH solution containing 25 mg ml-' of rabbit liver glycogen. The suspension was incubated at 45 C for 16 h with gentle shaking. The gel was then washed, successively, with 375 ml deionized H20, 750 ml of 25 mg/ml glucose solution, 375 ml deionized H20, 750 ml of 25 mg/ml glucose solution, 375 ml deionized H20, 375 ml of 0.1 M borate buffer (pH 8.0) containing 0.5 M KCI, 375 ml of 0.1 M Na-acetate buffer (pH 4.0) containing 0.5 M KCI, and finally with 375 ml deionized H20. The gel ...

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Section: Discussionmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

“…Recent studies indicate that multiple forms of phosphorylase do exist in spinach leaf (3,19,23). One of these forms appears to be localized within the chloroplast.…”

mentioning

confidence: 99%

See 2 more Smart Citations

Characterization of the Spinach Leaf Phosphorylases

Preiss¹,

Okita²,

Greenberg³

1980

Plant Physiol.

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show abstract

“…These two phosphorylases differ in affinity for various poly-and oligosaccharide substrates as well as molecular weight an(j some other enzymic properties. 5 ) Phosphorylaseisozymes were also detected in immature plants, .such as broad bean cotyledons 6 ) and pea cotyledons,7) where two types of activity were distinguished from the primer specificity for maltotriose. In the case of soybean, the occurrence of starch (0.2,,-,0.9% dry wt.)…”

Section: Received May 19 1986mentioning

confidence: 99%