Stable form of ascorbate peroxidase from the red alga Galdieria partita similar to both chloroplastic and cytosolic isoforms of higher plants

Depletion of the electron donor ascorbate causes rapid inactivation of chloroplastic ascorbate peroxidase (APX) of higher plants, while cytosolic APX is stable under such conditions. Here we report the cloning of cDNA from Galdieria partita, a unicellular red alga, encoding a novel type of APX (APX-...

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Bibliographic Details
Published in:Bioscience, biotechnology, and biochemistry biotechnology, and biochemistry, 2002-11, Vol.66 (11), p.2367-2375
Main Authors: Kitajima, S. (Research Inst. of Innovative Technology for the Earth, Kizu, Kyoto (Japan)), Ueda, M, Sano, S, Miyake, C, Kohchi, T, Tomizawa, K, Shigeoka, S, Yokota, A
Format: Article
Language:English
Subjects:
Amino Acid Sequence
Analytical, structural and metabolic biochemistry
APX-B gene
Ascorbate Peroxidases
ASCORBIC ACID
Ascorbic Acid - analogs & derivatives
Ascorbic Acid - metabolism
Base Sequence
Biological and medical sciences
cDNA
Chlorophyta - genetics
Chloroplasts - enzymology
Cytosol - enzymology
DNA, Complementary - genetics
Enzyme Stability
Enzymes
Enzymes and enzyme inhibitors
Fundamental and applied biological sciences. Psychology
Galdieria partita
Immunoblotting
inactivation
Isoenzymes - genetics
Isoenzymes - metabolism
Kinetics
Marine
Metabolism
Molecular Sequence Data
OXIDOREDUCTASES
Peroxidases - genetics
Peroxidases - metabolism
Plant physiology and development
Plant Proteins - genetics
Plant Proteins - metabolism
Plants - genetics
reactive oxygen species
recombinant protein
RECOMBINANT PROTEINS
Recombinant Proteins - genetics
Recombinant Proteins - metabolism
RHODOPHYCEAE
Rhodophyta
Rhodophyta - enzymology
Sequence Alignment
Sequence Homology, Amino Acid
Spectrophotometry
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Summary:Depletion of the electron donor ascorbate causes rapid inactivation of chloroplastic ascorbate peroxidase (APX) of higher plants, while cytosolic APX is stable under such conditions. Here we report the cloning of cDNA from Galdieria partita, a unicellular red alga, encoding a novel type of APX (APX-B). The electrophoretic mobility, K m values, k cat and absorption spectra of recombinant APX-B produced in Escherichia coli were measured. Recombinant APX-B remained active for at least 180 min after depletion of ascorbate. The amino-terminal half of APX-B, which forms the distal pocket of the active site, was richer in amino acid residues conserved in chloroplastic APXs of higher plants rather than cytosolic APXs. In contrast, the sequence of the carboxyl-terminal half, which forms the proximal pocket, was similar to that of the cytosolic isoform. The stability of APX-B might be due to its cytosolic isoform-like structure of the carboxyl-terminal half.
ISSN:0916-8451
1347-6947
DOI:10.1271/bbb.66.2367