Lipoic acid metabolism in Arabidopsis thaliana: Cloning and characterization of a cDNA encoding lipoyltransferase

Lipoic acid is an essential coenzyme required for activity of several key e nzyme 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 repor t the cloning and characterization of Arabidopsis thaliana LIP2 cDNA for li poyltransferase 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, roll mutant efficiently complemented the activity of pyruvate deh ydrogenase complex, but less efficiently than that of 2-oxoglutarate dehydr ogenase complex. Comparison of the deduced amino acid sequence of LIP2 with those of E, coli and yeast lipoyl-transferases showed a marked sequence si milarity and the presence of a leader sequence presumably required for impo rt 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 le aves. Western blot analysis with an antibody against lipoyltransferase demo nstrated that a 29 kDa form of lipoyltransferase is located in the mitochon drial compartment of A. thaliana.