Biotin-dependent carboxylases are ubiquitous enzymes necessary for a variety of anabolic and catabolic pathways. In plants, six biotinylated polypeptides have been detected (Wurtele and Nikolau, 1990), and four biotin-dependent carboxylase enzymes have been demonstrated: acetyl-COA carboxylase (Egli et al., 1993, and refs. therein), MCCase, pyruvate carboxylase, and propionyl-COA carboxylase (Wurtele and Nikolau, 1990). In mammals, MCCase is an enzyme of Leu degradation (Lau et al., 1980). The physiological function of MCCase in plants may be similar. We have isolated a full-length Arubidopsis cDNA clone of MCCase (BP-1), based on its ability to direct the synthesis of a biotinylated protein in Escherichia coli. This biotinylated protein presented epitopes that were recognized by antibodies directed against the biotinylated subunit of MC-Case from tomato (Wang et al., 1994). The deduced amino acid sequence derived from the nucleotide sequence of BP-1 showed high identity with the sequences of the biotinylated subunit of MCCase from tomato (Wang et al., 1994) and soybean (Song et al., 1994). These characterizations identify BP-1 as a clone coding for the biotinylated subunit of MCCase from Arabidopsis (Table I). Consistent with the mitochondrial location of MCCase in animals (Lau et al., 1980) and plants (Alban et al., 1993), the deduced amino terminus of the cDNA sequence has the characteristics of a mitochondrial transit peptide. A consensus mitochondrial cleavage site, RYIS, is found at residues 23 to 26 (von Heijne, 1992). If we assume that the RYI/S sequence is the junction between the transit peptide and the mature protein, then the mature protein of 690 amino acids has a calculated M, of 75,005. Based on sequence identity with biotin-containing carboxylases, the amino-terminal two-thirds of the MCCase biotinylated subunit represents the biotin carboxylase do-