Dehydroascorbate reductase cDNA from sweet potato (Ipomoea batatas [L.] Lam): expression, enzyme properties, and kinetic studies.
A cDNA encoding a putative dehydroascorbate reductase (DHAR) was cloned from sweet potato. The deduced protein showed a high level of sequence homology with DHARs from other plants (67 to approximately 81%). Functional sweet potato DHAR was overexpressed and purified. The purified enzyme showed an active monomeric form on a 12% native PAGE. The protein's half-life of deactivation at 50 degrees C was 10.1 min, and its thermal inactivation rate constant K(d) was 6.4 x 10(-2) min(-1). The enzyme was stable in a broad pH range from 6.0-11.0 and in the presence of 0.8 M imidazole. The K(m) values for DHA and GSH were 0.19 and 2.38 mM, respectively.
Jiang
Yu-Chi
YC
Institute of Bioscience and Biotechnology and Center for Marine Bioscience and Biotechnology, National Taiwan Ocean University, Keelung 202, Taiwan.
Huang
Chih-Yu
CY
Wen
Lisa
L
Lin
Chi-Tsai
CT
D000595:Amino Acid Sequence; D003001:Cloning, Molecular; D018076:DNA, Complementary / Q000235:genetics*; D015870:Gene Expression; D027723:Ipomoea batatas / Q000201:enzymology* / Q000235:genetics; D007700:Kinetics; D010088:Oxidoreductases / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism*; D011994:Recombinant Proteins / Q000235:genetics / Q000302:isolation & purification
D018076:DNA, Complementary; D011994:Recombinant Proteins; D010088:Oxidoreductases; C020666:glutathione dehydrogenase (ascorbate)