BIOCHEMICAL AND MOLECULAR BIOLOGICAL CHARACTERIZATION OF CAC2, THE ARABIDOPSIS-THALIANA GENE CODING FOR THE BIOTIN CARBOXYLASE SUBUNIT OF THE PLASTIDIC ACETYL-COENZYME-A CARBOXYLASE

The biotin carboxylase subunit of the heteromeric chloroplastic acetyl -coenzyme A carboxylase (ACCase) of Arabidopsis thaliana is coded by a single gene (CAC2), which is interrupted by 15 introns. The cDNA enco des a deduced protein of 537 amino acids with an apparent N-terminal c hloroplast-targeting transit peptide. Antibodies generated to a glutat hione S-transferase-CAC2 fusion protein react solely with a 51-kD poly peptide of Arabidopsis; these antibodies also inhibit ACCase activity in extracts of Arabidopsis. The entire CAC2 cDNA sequence was expresse d in Escherichia coli and the resulting recombinant biotin carboxylase was enzymatically active in carboxylating free biotin. The catalytic properties of the recombinant biotin carboxylase indicate that the act ivity of the heteromeric ACCase may be regulated by light-/dark-induce d changes in stromal pH. The CAC2 gene is maximally expressed in organ s and tissues that are actively synthesizing fatty acids for membrane lipids or oil deposition. The observed expression pattern of CAC2 mirr ors that previously reported for the CAC1 gene (J.-K. Choi, F. Yu, E.S . Wurtele, B.J. Nikolau [1995] Plant Physiol 109: 619-625;. Ke, J.-K. Choi, M. Smith, H.T. Horner, B.J. Nikolau, E.S. Wurtele [1997] Plant P hysiol 113: 357-365), which codes for the biotin carboxyl carrier subu nit of the heteromeric ACCase. This coordination is probably partially established by coordinate transcription of the two genes. This hypoth esis is consistent with the finding that the CAC2 and CAC1 gene promot ers share a common set of sequence motifs that may be important in gui ding the transcription of these genes.