Between the different types of Acyl-CoA dehydrogenases (ACADs), those specific for branched chain acyl-CoA derivatives are involved in the catabolism of amino acids. In mammals, isovaleryl-CoA dehydrogenase (IVD), an enzyme of the leucine catabolic pathway, is a mitochondrial protein, as other acyl-CoA dehydrogenases involved in fatty acid b-oxidation. In plants, fatty acid b-oxidation takes place mainly in peroxisomes, and the cellular location of the enzymes involved in the catabolism of branched-chain amino acids had not been definitely assigned.Here, we describe that highly purified potato mitochondria have important IVD activity. The enzyme was partially purified and cDNAs from two different genes were obtained. The partially purified enzyme has enzymatic constant values with respect to isovaleryl-CoA comparable to those of the mammalian enzyme. It is not active towards straight-chain acyl-CoA substrates tested, but significant activity was also found with isobutyryl-CoA, implying an additional role of the enzyme in the catabolism of valine. The present study confirms recent reports that in plants IVD activity resides in mitochondria and opens the way to a more detailed study of amino-acid catabolism in plant development.Keywords: plant mitochondria; isovaleryl-CoA dehydrogenase; leucine catabolism.The desaturation of the C 2 -C 3 bond of acyl-CoA esters to 2-trans-enoyl-CoA is an important step in the b-oxidation of fatty acids [1,2] and in the catabolism of branchedchain amino acids, leucine, isoleucine and valine [3,4]. In eukaryotes, this reaction can be catalyzed by two distinct type of enzymes: mitochondrial acyl-CoA dehydrogenases (ACADs), which feed reducing equivalents to the ubiquinone mitochondrial pool [5], and by peroxisomal acyl-CoA oxidases (ACOXs), which pass electrons directly on to molecular oxygen producing H 2 O 2 [2]. Both types of enzymes utilize FAD as cofactor. In mammalian tissues, b-oxidation takes place in both peroxisomes and mitochondria. In contrast, in higher plants this catabolism takes place mainly in peroxisomes [6±8], and the existence in higher plants of an additional mitochondrial b-oxidation system (and thus of ACAD enzymes) has often been considered controversial [9±11]. However, inhibition of mitochondrial b-oxidation by respiratory-chain inhibitors [9,12] strongly suggested that higher-plant mitochondria possess ACAD activities, which were identified in mitochondria from carbohydrate-starved maize root tips and sunflower embryos [12,13]. Moreover, putative plant ACAD genes could be identified on the basis of their sequence similarity with the corresponding mammalian enzymes [13]. However, a number of these putative ACADs have eventually been characterized as peroxisomal ACOXs [11,14].Among ACAD activities typically found in the mitochondria of animal cells, those specific for branched-chain substrates are involved in the catabolism of amino acids, in which the isobutyryl-CoA, 2-methyl-butyryl-CoA, and isovaleryl-CoA catabolites of Val, Ile, and Leu, respectively, und...

The gene and cDNA of an Arabidopsis thaliana cytidine deaminase (CDA) were cloned and sequenced. The gene, At-cda1, is located on chromosome 2 and is expressed in all plant tissues tested, although with quantitative differences. Expression analysis suggest that At-cda1 probably codes for the housekeeping cytidine deaminase of Arabidopsis. The gene was functionally expressed in Escherichia coli and the protein, At-CDA1, shows similar enzymatic and substrate specificities as conventional cytidine deaminases: it deaminates cytidine and deoxycytidine and is competitively inhibited by cytosine-containing compounds. Because the protein shows no affinity to RNA, it is not likely to be involved in RNA-editing by C-to-U deamination.When compared to cytidine deaminases from other organisms, it becomes clear that At-CDA1 is related, both in sequence and structure, to the CDA of E. coli and other gram-negative bacteria. The eubacterial nature of the Arabidopsis CDA suggests that it is an additional example of a plant gene of endosymbiotic origin.

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