PLANT PHYSIOLOGY , Vol 115, Issue 4 1371-1383, Copyright © 1997 by American Society of Plant Biologists
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BIOCHEMISTRY AND ENZYMOLOGY |
Biochemical and Molecular Biological Characterization of CAC2, the Arabidopsis thaliana Gene Coding for the Biotin Carboxylase Subunit of the Plastidic Acetyl-Coenzyme A Carboxylase
J. Sun, J. Ke, J. L. Johnson, B. J. Nikolau and E. S. Wurtele
Department of Botany (J.S., J.K., E.S.W.), and Department of Biochemistry and Biophysics (J.L.J., B.J.N.), Iowa State University, Ames, Iowa 50011-1020
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 encodes
a deduced protein of 537 amino acids with an apparent N-terminal
chloroplast-targeting transit peptide. Antibodies generated to a
glutathione S-transferase-CAC2 fusion protein react solely with a 51-kD
polypeptide of Arabidopsis; these antibodies also inhibit ACCase activity
in extracts of Arabidopsis. The entire CAC2 cDNA sequence was expressed 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 activity of the
heteromeric ACCase may be regulated by light-/dark-induced changes in
stromal pH. The CAC2 gene is maximally expressed in organs and tissues that
are actively synthesizing fatty acids for membrane lipids or oil
deposition. The observed expression pattern of CAC2 mirrors that previously
reported for the CAC1 gene (J.-K. Choi, F. Yu, E.S. Wurtele, B.J. Nikolau
[1995] Plant Physiol 109: 619-625; J. Ke, J.-K Choi, M. Smith, H.T. Horner,
B.J. Nikolau, E.S. Wurtele [1997] Plant Physiol 113: 357-365), which codes
for the biotin carboxyl carrier subunit of the heteromeric ACCase. This
coordination is probably partially established by coordinate transcription
of the two genes. This hypothesis is consistent with the finding that the
CAC2 and CAC1 gene promoters share a common set of sequence motifs that may
be important in guiding the transcription of these genes.
