Enzymes of the ascorbate biosynthesis and ascorbate-glutathione cycle in cultured cells of tobacco Bright Yellow 2

Pinto, M.C. de; Tommasi, F.; Gara, L. de

Plant Physiology and Biochemistry (Paris) 38(7-8): 541-550

2000


ISSN/ISBN: 0981-9428
DOI: 10.1016/s0981-9428(00)00773-7
Accession: 003434272

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Abstract
Ascorbate (ASC) and glutathione (GSH) metabolism were studied in cultured Nicotiana tabacum cv. Bright Yellow 2 (TBY-2) cells. TBY-2 cells contained L-galactono- gamma -lactone dehydrogenase (GLDH) (EC 1.3.2.3), an enzyme that converts L-galactono- gamma -lactone into ASC. Cellular fractionation of TBY-2 protoplasts indicated that this enzyme is exclusively localized in mitochondria and associated to the membrane fractions. During the growth cycle of TBY-2 cell culture, GLDH transiently increased, reaching the maximum value on the third day of culture, at the beginning of the exponential phase, when the cell proliferative activity was also higher. Similar behaviour has been observed for ASC and GSH contents. The activities of ascorbate peroxidase (APX) (EC 1.11.1.11), ascorbate-free radical reductase (AFRR) (EC 1.6.5.4), dehydroascorbic acid reductase (DHAR) (EC 1.8.5.1) and glutathione reductase (GR) (EC 1.6.4.2) also transiently increased. However, the scale of the increases varied being about 4-fold for APX and AFRR, 2-fold for DHAR and more than 11-fold for GR. The behaviour of the ASC and GSH recycling enzymes allowed TBY-2 cells to maintain both dehydroascorbic acid and glutathione disulphide at low levels, even under conditions of high ASC and GSH utilization. The relationship between the ASC and GSH metabolisms during the growth cycle of TBY-2 cell suspension cultures is also discussed.