Isolation and Characterization of Vacuoles from Cell Suspension Cultures of Daucus carota

1979 ◽  
Vol 34 (9-10) ◽  
pp. 848-853 ◽  
Author(s):  
Florenz Sasse ◽  
Dietlinde Backs-Hüsemann ◽  
Wolfgang Barz
Keyword(s):  
Amino Acids ◽  
Cell Suspension ◽  
Daucus Carota ◽  
Osmotic Shock ◽  
Cell Suspension Cultures ◽  
Large Data ◽  
Total Activity ◽  
Suspension Cultures ◽  
Anthocyanin Content ◽  
Isolation And Characterization

Abstract A reliable procedure for the isolation of vacuoles from anthocyanin-containing cells of Daucus carota cell suspension cultures has been developed. From cells of the late linear growth phase, protoplasts were prepared and purified in a sucrose/sorbitol gradient. Vacuoles were liberated from these protoplasts by osmotic shock and purified in a Metrizamide step gradient. The vacuolecontaining fractions were analysed for their anthocyanin content as a measure for the yield of vacuoles. The purity of vacuoles was examined by assaying various marker enzymes in both protoplasts and vacuoles. The purest vacuolar fraction had 8% of the total activity of glucose-6-phosphate dehydrogenase (marker for cytosol), 8% of cytochrome oxidase (mitochondria) and 10% of NADPH -cytochrome c reductase (ER). 55% of the acid phosphatase activity of the protoplasts and 35% of the total malate were recovered in the vacuoles. The vacuolar pool of amino acids is quite large. Data for 15 amino acids show that 44 to 73% are being located in the vacuole.

Isolation and Characterization of Flavonol Converting Enzymes from Mentha piperita Plants and from Mentha arvensis Cell Suspension Cultures

1979 ◽  
Vol 34 (3-4) ◽  
pp. 200-209 ◽  
Author(s):  
Gudrun Frey-Schröder ◽  
Wolfgang Barz
Keyword(s):  
Cell Suspension ◽  
Oxidase Activity ◽  
Enzyme Reaction ◽  
Cell Suspension Cultures ◽  
Suspension Cultures ◽  
Mentha Piperita ◽  
Mentha Arvensis ◽  
Iaa Oxidase ◽  
Isolation And Characterization ◽  
Group I

Abstract Peroxidases from several plants, including horseradish peroxidase, were capable of converting flavonols to the corresponding 2,3-dihydroxyflavanones in presence of H2O2 . Contrastingly, protein extracts from Mentha piperita plants and Mentha arvensis cell suspension cultures perform ed the same enzymatic step in absence of H2O2 , but only with quercetin, not with kaempferol. H2O2-independent, quercetin converting enzymes were isolated and purified from these extracts, and they could be classified in two groups according to the extent of stimulation of the enzyme reaction by H2O2 . Enzymes from group I were stimulated by exogenous H2O2 , and they resembled horse­ radish peroxidase in several aspects. They possessed IAA oxidase activity, but quercetin was the preferred substrate. Enzymes from group II from the plants appeared to be a distinctly different set of enzymes. They were not stimulated by H2O2 , but required molecular oxygen and converted only 3,3′,4′-trihydroxyflavones under aerobic conditions. Also, they showed no Soret-bands and possessed no IAA oxidase activity. These proteins appear to be a new class of enzymes participating in the first step of flavonol degradation in plants.

Isolation and characterization of an UDPG-dependent glucosyltransferase activity from Rauwolfia serpentina Benth. cell suspension cultures

1994 ◽  
Vol 72 (1) ◽  
pp. 51-55 ◽  
Author(s):  
Ralf Lutterbach ◽  
Carl Michael Ruyter ◽  
Joachim Stöckigt
Keyword(s):  
Enzyme Activity ◽  
Cell Suspension ◽  
Cell Suspension Cultures ◽  
Callus Cultures ◽  
Suspension Cultures ◽  
Rauwolfia Serpentina ◽  
Isolation And Characterization ◽  
Membrane Bound ◽  
Plant Families

From cell suspension cultures of Rauwolfia serpentina Benth. a new enzyme activity was isolated and its properties determined. The enzyme is a soluble protein and catalyzes the transfer of a glucose moiety from UDPG to a wide variety of phenolic compounds with p-nitrophenol as one of the best substrates (Km = 1.21 mM, UDPG = 0.54 mM). In contrast to the membrane-bound UDPG: vomilenine-21-OH-β-D-glucosyltransferase from Rauwolfia serpentina cells, this enzyme is not able to glucosylate indole alkaloids. The enzyme activity has been detected in 14 callus cultures belonging to 10 different plant families.

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