J. Biol. Chem., Vol. 268, Issue 4, 2363-2367, 02, 1993
Import and assembly of the beta-subunit of chloroplast coupling factor 1 (CF1) into isolated intact chloroplasts
GG Chen and AT Jagendorf
Section of Plant Biology, Cornell University, Ithaca, New York 14853.
The transit peptide gene of the small subunit of ribulose-1,5- bisphosphate
carboxylase/oxygenase from Nicotiana plumbaginifolia was fused to the
tentoxin-resistant beta-subunit gene of chloroplast coupling factor 1 (CF1)
from Nicotiana tabacum via a linker sequence. The consequent fusion gene
encodes the entire gene sequences of both the transit peptide and the
beta-subunit of CF1 without a single change of amino acid residues. The
fusion gene was in vitro expressed in a coupled transcription-translation
system as a 62-kDa precursor and was imported into isolated intact
chloroplasts of Nicotiana longiflora. The imported precursor was found to
be processed to the expected mature beta-subunit size. Evidence is
presented that radioactively labeled beta-subunit was incorporated into
mature CF1 and not just nonspecifically associated with the thylakoid
membranes. Since the fusion protein containing only the transit peptide
sequence was imported and apparently correctly processed, it may not be
necessary to include N-terminal amino acids of the mature small subunit for
correct proteolytic cleavage in the chloroplast stroma.
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