Targeting of proteins to the thylakoid lumen by the bipartite transit peptide of the 33 kd oxygen-evolving protein.
Various chimeric precursors and deletions of the 33 kd oxygen-evolving protein (OEE1) were constructed to study the mechanism by which chloroplast proteins are imported and targeted to the thylakoid lumen. The native OEE1 precursor was imported into isolated chloroplasts, processed and localized in the thylakoid lumen. Replacement of the OEE1 transit peptide with the transit peptide of the small subunit of ribulose-1,5-bisphosphate carboxylase, a stromal protein, resulted in redirection of mature OEE1 into the stromal compartment of the chloroplast. Utilizing chimeric transit peptides and block deletions we demonstrated that the 85 residue OEE1 transit peptide contains separate signal domains for importing and targeting the thylakoid lumen. The importing domain, which mediates translocation across the two membranes of the chloroplast envelope, is present in the N-terminal 58 amino acids. The thylakoid lumen targeting domain, which mediates translocation across the thylakoid membrane, is located within the C-terminal 27 residues of the OEE1 transit peptide. Chimeric precursors were constructed and used in in vitro import experiments to demonstrate that the OEE1 transit peptide is capable of importing and targeting foreign proteins to the thylakoid lumen.
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Department of Biology, University of Pennsylvania, Philadelphia 19104.
Cashmore
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D020418:Algal Proteins; D000595:Amino Acid Sequence; D001692:Biological Transport; D002678:Chimera / Q000235:genetics; D002736:Chloroplasts / Q000378:metabolism*; D003001:Cloning, Molecular; D004247:DNA / Q000235:genetics; D017343:Genes, Plant; D008969:Molecular Sequence Data; D010455:Peptides / Q000378:metabolism*; D010940:Plant Proteins / Q000235:genetics / Q000378:metabolism*; D010957:Plasmids; D011498:Protein Precursors / Q000235:genetics / Q000378:metabolism; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D015183:Restriction Mapping; D012273:Ribulose-Bisphosphate Carboxylase / Q000235:genetics / Q000378:metabolism; D014158:Transcription, Genetic
D020418:Algal Proteins; D010455:Peptides; D010940:Plant Proteins; D011498:Protein Precursors; D011993:Recombinant Fusion Proteins; C052389:oxygen-evolving enhancer protein 1, plant; D004247:DNA; D012273:Ribulose-Bisphosphate Carboxylase