IMPORT OF BARLEY PHOTOSYSTEM-I SUBUNIT-N INTO THE THYLAKOID LUMEN IS MEDIATED BY A BIPARTITE PRESEQUENCE LACKING AN INTERMEDIATE PROCESSINGSITE - ROLE OF THE DELTA-PH IN TRANSLOCATION ACROSS THE THYLAKOID MEMBRANE

Translocation across the thylakoid membrane of the recently identified photosystem I polypeptide, PSI-N, has been analyzed in pea (Pisum sat ivum) and barley (Hordeum vulgare). PSI-N from barley is synthesized i n the cytosol with a bipartite presequence similar in structural terms to those of other cytosolically synthesized proteins routed to the th ylakoid lumen. In vitro reconstitution assays demonstrate that translo cation into thylakoids is absolutely dependent on the trans-thylakoida l DELTApH, but that nucleotide triphosphates are not required; the tra nslocation mechanism is thus similar in these respects to those utiliz ed by the 23- and 16-kDa proteins of the oxygen-evolving complex. The translocation of PSI-N is unique in that the presequence of PSI-N does not contain an intermediate cleavage site for the stromal processing peptidase; import experiments using intact chloroplasts depleted of a DELTApH by nigericin treatment demonstrate the accumulation of the ful l precursor protein in the stroma. Translocation across the thylakoid membrane can take place in the absence of stromal factors, although th e presence of stromal extracts leads to a consistent but slight enhanc ement of translocation efficiency. We also show that efficient translo cation of the 33-kDa protein of the oxygen-evolving complex can take p lace in the complete absence of DELTApH, in apparent contradiction wit h earlier findings; the translocation of this protein is thus similar in several respects to that of plastocyanin. The data indicate the ope ration of two very different types of translocation mechanism, with PS I-N exhibiting additional separate characteristics.