Reconstitution of barley photosystem I reveals that the N-terminus of the PSI-D subunit is essential for tight binding of PSI-C |
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Authors: | Helle Naver M Paul Scott Birgitte Andersen Birger Lindberg Møller Henrik Vibe Scheller |
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Institution: | Dept of Plant Biolgy. The Royal Veterinary and Agricultural Univ., 40 Thorvaldsensvej. DK-1871 Frederiksberg C, Copenhagen, Denmark. |
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Abstract: | Removal of the peripheral subunits PSI-C, -D and -E from the photosystem I (PSI) complex of barley requires a urea treatment much harsher than required to remove the similar subunits from cyanobacterial PSI. The resulting PSI barley core was reconstituted by addition of the E. coli expressed subunits PSI-C and -D, and PSI-E isolated from barley. Western blotting, flash photolysis and NADP+ photoreduction measurements demonstrated complete and specific removal of the three subunits from the core and efficient reconstitution of the complex after addition of PSI-C, -D and -E. Flash photolysis reveals that PSI-D is essential for binding of functional PSI-C to the PSI core. An N-terminally truncated barley PSI-D lacking 24 amino acid residues and thus being without the N-terminal extension characteristic for higher plant PSI-D proteins reconstitutes the PSI core to 50% of the level obtained with intact PSI-D as demonstrated by flash photolysis and NADP+ photoreduction measurements. Cyanobacterial PSI-D is functionally equivalent to truncated barley PSI-D with respect to its activity to reconstitute the PSI core. This shows that the N-terminal extension of plant PSI-D plays a key role in binding PSI-C to the core. The plant-specific N-terminus of PSI-D is hypothesized to execute its function through interaction with a plant-specific PSI subunit, possibly PSI-H. An anchoring function of the N-terminus of PSI-D would also explain the harsh treatment needed to obtain a plant PSI core. PSI-E is important for efficient NADP+ reduction but does not influence electron transfer to iron-sulphur centres A/B nor binding of PSI-C. The enhancing effect of PSI-E on NADP+ reduction is independent of the presence of the N-terminus of PSI-D. |
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Keywords: | Barley flash photolysis Hordeum vulgare photosystem I PSI-C PSI-D PSI-E reconstitution |
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