Acyl carrier protein (ACP) import into chloroplasts. Covalent modification by a stromal holoACP synthase is stimulated by exogenously added CoA and inhibited by adenosine 3',5'-bisphosphate.
During the import of the precursor for the acyl carrier protein (ACP) into chloroplasts, apoACP is converted to holoACP by the attachment of a phosphopantetheine group transferred from coenzyme A (CoA) by a chloroplast holoACP synthase [Fernandez, M. and Lamppa, G. (1990) Acyl carrier protein import into chloroplasts does not require the phosphopantetheine: evidence for a chloroplast holoACP synthase, Plant Cell 2, 195-206]. Here it is shown that exogenous addition of CoA to intact chloroplasts in the import assay stimulates the conversion of apoACP to holoACP. If adenosine 3',5'-bisphosphate [Ado(3',5')P2], the byproduct of the transfer reaction, was also included the extent of conversion was greatly reduced. CoA has its effect after ACP precursor (pre-ACP) import and proteolytic removal of the transit peptide, thus indicating that the chloroplast holoACP synthase resides in the stroma where fatty acid synthase is found. When Ado(3',5')P2 was added alone to the import assay, it inhibited the synthesis of holoACP. Inhibition of the conversion of apo- to holoACP with Ado(3',5')P2 made it possible to examine whether the holoform of preACP could be imported into chloroplasts. Pre-apoACP was synthesized in Escherichia coli and shown to be competent for import in an ATP- and temperature-dependent manner. A partially purified chloroplast holoACP synthase converted 60-90% of the pre-apoACP to pre-holoACP. Pre-holoACP incubated with chloroplasts in the presence of Ado(3',5')P2 yielded > 60% holoACP, whereas the control reaction with pre-apoACP gave primarily apoACP. Hence the phosphopantetheine prosthetic group of ACP does not block precursor movement through the translocation apparatus of the chloroplast envelope.
Yang
L M
LM
Department of Molecular Genetics and Cell Biology, University of Chicago, IL 60637.
Fernandez
M D
MD
Lamppa
G K
GK
D000213:Acyl Carrier Protein / Q000096:biosynthesis*; D000244:Adenosine Diphosphate / Q000494:pharmacology*; D001059:Apoproteins / Q000378:metabolism*; D002736:Chloroplasts / Q000378:metabolism*; D003065:Coenzyme A / Q000494:pharmacology*; D004926:Escherichia coli; D007700:Kinetics; D015388:Organelles / Q000378:metabolism; D011499:Protein Processing, Post-Translational; D011994:Recombinant Proteins / Q000378:metabolism; D018724:Spinacia oleracea / Q000378:metabolism; D017855:Transferases (Other Substituted Phosphate Groups)
D000213:Acyl Carrier Protein; D001059:Apoproteins; D011994:Recombinant Proteins; D000244:Adenosine Diphosphate; C010840:adenosine 3'-phosphate-5'-phosphate; D017855:Transferases (Other Substituted Phosphate Groups); C021364:holo-(acyl-carrier-protein) synthase; D003065:Coenzyme A