Sec-dependent pathway and DeltapH-dependent pathway do not share a common translocation pore in thylakoidal protein transport.
Thylakoidal proteins of plant chloroplasts are transported to thylakoids via several different pathways, including the DeltapH-dependent and the Sec-dependent pathways. In this study, we asked if these two pathways utilize a common translocation pore. A fusion protein consisting of a 23-kDa subunit of the oxygen evolving complex and Escherichia coli biotin carboxyl carrier protein was biotinylated in E. coli cells and purified. When incubated with isolated pea thylakoids in the absence of avidin, the purified fusion protein was imported into the thylakoids via the DeltapH-dependent pathway. However in the presence of avidin, the fusion protein became lodged in the thylakoid membranes, with its N terminus reaching the thylakoidal lumen, while its C-terminal segment complexed with avidin exposed on the thylakoidal surface. The translocation intermediate of the fusion protein inhibited the import of authentic 23-kDa subunit, suggesting that it occupies a putative translocation pore for the DeltapH-dependent pathway. However the intermediate did not block import of the 33-kDa subunit of the oxygen evolving complex, which is a substrate for the Sec-dependent pathway. These results provide evidence against the possibility of a common translocation pore shared by the Sec-dependent pathway and the DeltapH-dependent pathway.
Department of Chemistry, Faculty of Science, Nagoya University, Chikusa-ku, Nagoya 464-8602, Japan.
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T
Asai
Y
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Shinoda
T
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Nohara
T
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Yoshihisa
T
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Endo
D000103:Acetyl-CoA Carboxylase / Q000378:metabolism; D001360:Avidin / Q000378:metabolism; D001426:Bacterial Proteins / Q000378:metabolism*; D001692:Biological Transport; D001710:Biotin / Q000378:metabolism; D002352:Carrier Proteins / Q000378:metabolism; D054889:Fatty Acid Synthase, Type II; D006863:Hydrogen-Ion Concentration; D006868:Hydrolysis; D007425:Intracellular Membranes / Q000378:metabolism; D021382:Protein Sorting Signals / Q000378:metabolism
D001426:Bacterial Proteins; D002352:Carrier Proteins; D021382:Protein Sorting Signals; D001360:Avidin; D001710:Biotin; D054889:Fatty Acid Synthase, Type II; D000103:Acetyl-CoA Carboxylase; C079633:biotin carboxyl carrier protein