Journal Article

Lipoic Acid Metabolism in <i>Arabidopsis thaliana</i>: Cloning and Characterization of a cDNA Encoding Lipoyltransferase

Miho Wada, Rie Yasuno, Sean W. Jordan, John E. Cronan Jr. and Hajime Wada

in Plant and Cell Physiology

Published on behalf of Japanese Society of Plant Physiologists

Volume 42, issue 6, pages 650-656
Published in print June 2001 | ISSN: 0032-0781
Published online June 2001 | e-ISSN: 1471-9053 | DOI: http://dx.doi.org/10.1093/pcp/pce081
Lipoic Acid Metabolism in Arabidopsis thaliana: Cloning and Characterization of a cDNA Encoding Lipoyltransferase

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Lipoic acid is an essential coenzyme required for activity of several key enzyme complexes, such as the pyruvate dehydrogenase complex, in the central metabolism. In these complexes, lipoic acid must be covalently attached to one of the component proteins for it to have biological activity. We report the cloning and characterization of Arabidopsis thaliana LIP2 cDNA for lipoyltransferase that catalyzes the transfer of the lipoyl group from lipoyl-acyl carrier protein to lipoate-dependent enzymes. This cDNA was shown to code for lipoyltransferase by its ability to complement an Escherichia coli lipB null mutant lacking lipoyltransferase activity. The expressed enzyme in the E. coli mutant efficiently complemented the activity of pyruvate dehydrogenase complex, but less efficiently than that of 2-oxoglutarate dehydrogenase complex. Comparison of the deduced amino acid sequence of LIP2 with those of E. coli and yeast lipoyltransferases showed a marked sequence similarity and the presence of a leader sequence presumably required for import into mitochondria. Southern and northern hybridization analyses suggest that LIP2 is a single-copy gene and is expressed as an mRNA of 860 nt in leaves. Western blot analysis with an antibody against lipoyltransferase demonstrated that a 29 kDa form of lipoyltransferase is located in the mitochondrial compartment of A. thaliana.

Keywords: Key words: Arabidopsis thaliana — Fatty acid synthesis — Lipoic acid — Lipoyltransferase — Pyruvate dehydrogenase.; Abbreviations: ACP, acyl carrier protein; IPTG, isopropyl-1-thio-β-d-galactoside; OGDH, 2-oxoglutarate dehydrogenase; ORF, open reading frame; PDH, pyruvate dehydrogenase; 5′-RACE, 5′-rapid amplification of cDNA ends.

Journal Article.  5457 words.  Illustrated.

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

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