4.7 Article

Isolation and expression of protein disulfide isomerase cDNA from sweet potato (Ipomoea batatas [L.] Lam 'Tainong 57') storage roots

Journal

PLANT SCIENCE
Volume 169, Issue 4, Pages 776-784

Publisher

ELSEVIER IRELAND LTD
DOI: 10.1016/j.plantsci.2005.05.034

Keywords

sweet potato; protein disulfide isomerase; cDNA sequence; expression; dehydroascorbate; monodehydroascorbate

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A protein disulfide isomerase (PDI) coding sequence was cloned by differential display from sweet potato (Ipomoea batatas [L.] Lam 'Tainong 57') storage roots. The cDNA contains 1869 bp in length and encodes a protein of 503 amino acids. Primary structure analysis of the deduced protein revealed two thioredoxin-like active sites (CGHC), and an endoplasmic reticulum-retention signal at its C-terminus (KDEL), which is also found in PDIs of plants and animals. Although there is only about 65% amino acid identity between SPPD11 and other plant PDIs, the active site regions are almost identical. A recombinant protein expressed from the storage root cDNA clone effectively catalyzed glutathione-insulin transhydrogenation. SPPD11 reduced dehydroascorbate (DHA) in the presence of glutathione to regenerate L-ascorbic acid (AsA). However, without glutathione, SPPD11 has very low DHA reductase activity. And AsA was oxidized by AsA oxidase to generate monodehydroascorbate (MDA) free radical. MDA was also reduced by SPPD11 to AsA in the presence of NADH mimicking the MDA reductase catalyzed reaction. These data suggest that SPPD11 have both DHA reductase and MDA reductase activities. The corresponding mRNA level was found the highest in the storage roots, followed by sprouts and full-expanded green leaves; while was the lowest in sprouted roots and vein. In Western blot analysis, SPPD11 level was the highest in the storage roots, followed by sprouts, full-expanded green leaves and vein and no signal at all in sprouted root. These results suggested that the protein coded for by the sweet potato gene is a novel member of the PDI family in plants. SPPDI genes of sweet potato storage roots display differential gene expression patterns, which may be associated with the antioxidant functions they play in plant physiology in order to cope with particular developmental cue. (c) 2005 Elsevier Ireland Ltd. All rights reserved.

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