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In vitro study of the xanthine dehydrogenase from illuminated or darkened leaves
Author(s) -
Nguyen J.,
Nato A.
Publication year - 1983
Publication title -
physiologia plantarum
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.351
H-Index - 146
eISSN - 1399-3054
pISSN - 0031-9317
DOI - 10.1111/j.1399-3054.1983.tb06573.x
Subject(s) - hypoxanthine , nicotiana tabacum , biochemistry , enzyme assay , xanthine oxidase , enzyme , xanthine dehydrogenase , dehydrogenase , cycloheximide , xanthine , purine , darkness , biology , chemistry , botany , protein biosynthesis , gene
The activity of xanthine dehydrogenase (XDH; EC 1.2.3.37), a key enzyme of purine catabolism leading to the synthesis of ureides, was investigated in cell‐free extracts of illuminated or darkened tobacco ( Nicotiana tabacum L. cv. xanthi) leaves. Whereas the in vivo enzyme activity measured on leaf discs was lower in light than in darkness, cell‐free extracts from illuminated or darkened leaves exhibited the same maximum velocity around 30 pmol (mg fresh weight) ‐1 h ‐2 . The kinetic constants of the enzyme for the substrate and for the cofactor did not change in relation to illumination of leaves, the K m for hypoxanthine being about 30 μM and that for NAD + 13 μM. Extracts run on polyacrylamide gel electrophoresis showed only one band of XDH activity which was identical whether the plants were illuminated or not. The molecular weight of the native enzyme from both extracts was estimated to be 320 000 dalton. Dithiothreitol treatment of leaf extracts could not mimick the effect of light on XDH activity in leaves. A direct effect of light on the activity of that flavoprotein could not be observed specifically in relation to the light treatment of cotyledons of Pharbitis nil prior to the enzyme extraction. A cycloheximide‐trealment of cotyledons of Pharbitis nil decreased the in vivo XDH activity. But the inhibition of the enzyme activity was similar whether the plants were in conditions inducing an increase of activity or not, indicating that quantitative changes of the enzyme protein are not involved in the variations of the in vivo enzyme activity. The results are discussed in relation to the conditions prevailing in the in vitro and the in vivo enzyme activity measurements. It is suggested that light might affect the XDH activity in leaves through the modifications of cell environment.