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Mitochondrial and peroxisomal ascorbate peroxidase of pea leaves
Author(s) -
Jiménez A.,
Hernández J. A.,
Barceló A. Ros,
Sandalio L. M.,
Del Río L. A.,
Sevilla F.
Publication year - 1998
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 - 10.1034/j.1399-3054.1998.1040424.x
Subject(s) - apx , peroxidase , pyrogallol , peroxisome , biochemistry , chemistry , enzyme , gene
The isoenzyme pattern and the substrate specificity of the membrane‐bound mitochondrial and peroxisomal ascorbate peroxidases (APX; EC 1.11.1.11) from pea leaves are studied. The substrate specificity of both APXs was assayed using the electron donors ascorbate and pyrogallol, whereas o ‐dianisidine, hydroquinone, tetramethylbenzidine and 4‐methoxy‐α‐naphthol were also assayed with mitochondrial APX (mitAPX). In leaf mitochondria, the specific activity of APX was similar with pyrogallol and ascorbate, the activity being inhibited by p ‐CMS. mitAPX showed low activity with the guaiacol peroxidase (GPX)‐type substrates, tetramethylbenzidine and 4‐methoxy‐α‐naphthol. Activity of mitAPX with hydroquinone suggest a potential role of mitAPX in the drainage of electrons from the mitochondrial electron chain at the level of ubiquinone. In peroxisomes, the APX (perAPX) specific activity was much higher with pyrogallol than with ascorbate. This perAPX was more sensitive to incubation with Triton X‐100 than the mitAPX. By native PAGE the mitAPX was resolved in 6 isoenzyme bands, and the activity of the 3 main bands (mitAPX III, III′ and IV) was inhibited by p ‐CMS. These 3 major isozymes were also present in mitochondrial membrane fractions. Staining for GPX activity with 4‐methoxy‐α‐naphthol revealed that the APX detected in mitochondria did not have the capacity to oxidize 4‐MN, and therefore cannot be considered as true GPX. When intact peroxisomes and peroxisomal membranes were subjected to native PAGE, no APX activity could be detected and this was probably due to the inactivation of perAPX. Results obtained suggest that pea mitochondrial APX (mitAPX) represent a distinct and novel isozyme different from those APXs of chloroplast and cytosolic origin previously reported. The peroxisomal APX (perAPX), however, appears to ressemble the chloroplast APXs as regards its sensitivity to Triton X‐100.