Partial purification and characterization of a copper-induced anionic peroxidase of sunflower roots.
Treatment of 14-day-old sunflower seedlings with a toxic amount of copper (50 microM of CuSO(4)) during 5days caused significant increase in peroxidase activity in roots. Qualitative analysis of soluble proteins using native anionic PAGE followed by detection of peroxidase activity with guaïacol as electron donor in the presence of H(2)O(2) revealed five stimulated peroxidases, named A1, A2, A3, A4, and A5. These peroxidases had differential behavior during the period of treatment. A1, A2, A3 and A4 were stimulated in the first period of stress, but rapidly suppressed at 72h. A5 showed a progressive stimulation which was even increased at 120h. A1 was partially purified, identified using liquid chromatography coupled to mass spectrometry (LC-MS/MS), and characterized. Effects of pH and temperature on its activity were determined with guaïacol as electron donor. Optima were obtained at pH 8 and at 40 degrees C. Analysis of substrate specificity showed that A1 was active on coniferyl alcohol but not on IAA. Enzymatic activity was inhibited by a high concentration of H(2)O(2).
Jouili
Hager
H
Faculté des Sciences de Bizerte, Laboratoire de Biologie et Physiologie Cellulaires, 7021 Zarzouna, Bizerte, Tunisia.
Bouazizi
Houda
H
Rossignol
Michel
M
Borderies
Gisèle
G
Jamet
Elisabeth
E
El Ferjani
Ezzeddine
E
D000595:Amino Acid Sequence; D002853:Chromatography, Liquid; D019327:Copper Sulfate / Q000378:metabolism / Q000633:toxicity*; D004591:Electrophoresis, Polyacrylamide Gel; D006139:Guaiacol / Q000378:metabolism; D006368:Helianthus / Q000187:drug effects / Q000201:enzymology* / Q000378:metabolism; D006861:Hydrogen Peroxide / Q000378:metabolism; D006863:Hydrogen-Ion Concentration; D008969:Molecular Sequence Data; D010544:Peroxidases / Q000096:biosynthesis / Q000302:isolation & purification*; D018517:Plant Roots / Q000187:drug effects / Q000201:enzymology* / Q000378:metabolism; D036226:Seedlings / Q000187:drug effects / Q000201:enzymology / Q000378:metabolism; D016415:Sequence Alignment; D013379:Substrate Specificity; D053719:Tandem Mass Spectrometry; D013696:Temperature
D006139:Guaiacol; D006861:Hydrogen Peroxide; D010544:Peroxidases; C120494:anionic peroxidase; D019327:Copper Sulfate