Journal Article

Molecular Characterization of Tobacco Mitochondrial <span class="smallCaps">l</span>-Galactono-<i>γ</i>- Lactone Dehydrogenase and Its Expression in <i>Escherichia coli</i>

Yukinori Yabuta, Kazuya Yoshimura, Toru Takeda and Shigeru Shigeoka

in Plant and Cell Physiology

Published on behalf of Japanese Society of Plant Physiologists

Volume 41, issue 6, pages 666-675
Published in print June 2000 | ISSN: 0032-0781
Published online June 2000 | e-ISSN: 1471-9053 | DOI:
Molecular Characterization of Tobacco Mitochondrial l-Galactono-γ- Lactone Dehydrogenase and Its Expression in Escherichia coli

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  • Biochemistry
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  • Plant Sciences and Forestry


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A cDNA clone encoding l-galactono-ρ-lactone (GAL) dehydrogenase (EC was isolated from tobacco leaves. The cDNA clone contained an open reading frame encoding the protein of 501 amino acids with a calculated molecular mass of 56,926 Da, preceded by a putative mitochondrial targeting signal consisting of 86 amino acid residues. In fact, GAL dehydrogenase was localized in the mitochondria of tobacco cells. The deduced amino acid sequence of the cDNA showed 77 and 82% homology to cauliflower and sweet potato GAL dehydrogenases, respectively. Southern blot analysis showed that tobacco contains one copy of the gene for the enzyme. Northern blot analysis showed that GAL dehydrogenase mRNA (2.0 kb) is expressed in the leaves, stems, and roots in almost equal quantities. We introduced the cDNA clone encoding tobacco GAL dehydrogenase into a pET expression vector to overexpress this protein in Escherichia coli. The partially purified recombinant enzyme was used for comparative studies on the native enzymes from tobacco and other sources; its enzymatic properties were similar to those of other GAL dehydrogenases.

Keywords: Ascorbate biosynthesis; l-galactono-ρ-lactone dehydrogenase (EC; Nucleotide sequence; Recombinant enzyme; Tobacco (Nicotiana tabacum cv. Xanthi)

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Subjects: Biochemistry ; Molecular and Cell Biology ; Plant Sciences and Forestry

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