Characterization of ascorbate peroxidases from unicellular red alga Galdieria partita.
Galdieria partita, a unicellular red alga isolated from acidic hot springs and tolerant to sulfur dioxide, has at least two ascorbate peroxidase (APX) isozymes. This was the first report to demonstrate that two isozymes of APX are found in algal cells. Two isozymes were separated from each other at the hydrophobic chromatography step of purification and named APX-A and APX-B after the elution order in the chromatography. APX-B accounted for 85% of the total activity. Both isozymes were purified. APXs from Galdieria were monomers whose molecular weights were about 28,000, similar to stromal APX of higher plants. APX-A cross-reacted with monoclonal antibody raised against APX of Euglena gracilis in immunoblotting, but APX-B did not, although the antibody can recognize all other APXs tested. The amino-terminal sequences of APX-A and -B from Galdieria had some homology with each other but little homology with those from other sources. Their Km values for ascorbate and hydrogen peroxide were comparable with those of APX from higher plants. Unlike the green algal enzymes, the donor specificities of Galdieria APXs were as high as those of plant chloroplastic APX. On the contrary, these APXs reduced tertiary-butyl hydroperoxide as an electron acceptor as APXs from Euglena and freshwater Chlamydomonas do. The inhibition of APX-A and -B by cyanide and azide, and characteristics of their light absorbance spectra indicated that they were heme peroxidases.
Sano
S
S
Research Institute of Innovative Technology for the Earth, Kizu, Kyoto, 619-0292 Japan.
Ueda
M
M
Kitajima
S
S
Takeda
T
T
Shigeoka
S
S
Kurano
N
N
Miyachi
S
S
Miyake
C
C
Yokota
A
A
D000595:Amino Acid Sequence; D060387:Ascorbate Peroxidases; D004791:Enzyme Inhibitors / Q000494:pharmacology; D004795:Enzyme Stability; D006358:Hot Temperature; D007527:Isoenzymes / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism; D007700:Kinetics; D008969:Molecular Sequence Data; D008970:Molecular Weight; D010446:Peptide Fragments / Q000737:chemistry / Q000302:isolation & purification; D010544:Peroxidases / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism*; D000461:Rhodophyta / Q000201:enzymology*; D016415:Sequence Alignment; D017386:Sequence Homology, Amino Acid; D013053:Spectrophotometry; D018724:Spinacia oleracea / Q000201:enzymology; D013816:Thermodynamics
D004791:Enzyme Inhibitors; D007527:Isoenzymes; D010446:Peptide Fragments; D010544:Peroxidases; D060387:Ascorbate Peroxidases