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J Agric Food Chem. 2009 May 27;57(10):4403-8. doi: 10.1021/jf900045p. Epub 2009 Apr 9.

Cloning, expression, and purification of a functional glutathione reductase from sweet potato (Ipomoea batatas [L.] Lam): kinetic studies and characterization.

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Institute of Bioscience and Biotechnology and Center for Marine Bioscience and Biotechnology, National Taiwan Ocean University, Keelung 202, Taiwan.


A cDNA encoding a putative glutathione reductase (GR) was cloned from sweet potato (Ib). The deduced protein showed high level of sequence homology with GRs from other plants (79-38%). A three-dimensional (3-D) homology structure was created. The active site Cys residues are conserved in all reported GR. Functional IbGR was overexpressed and purified. The purified enzyme showed an active monomeric form on a 10% native polyacrylamide gel electrophoresis (PAGE). The monomeric nature of the enzyme was confirmed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and molecular mass determination of the native enzyme. The Michaelis constant (K(m)) values for GSSG (glutathione disulfide) and NADPH (β-nicotinamide adenine dinucleotide phosphate, reduced form) were 0.114 and 0.056 mM, respectively. The enzyme activity was inhibited by Cu(2+) and Zn(2+), but not by Ca(2+). The protein's half-life of deactivation at 70 °C was 3.3 min, and its thermal inactivation rate constant K(d) was 3.48 × 10(-1) min(-1). The enzyme was active in a broad pH range from 6.0 to 11.0 and in the presence of imidazole up to 0.8 M. The native enzyme appeared to be resistant to digestion by trypsin or chymotrypsin.

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