RECONSTITUTION OF BARLEY PHOTOSYSTEM-I REVEALS THAT THE N-TERMINUS OFTHE PSI-D SUBUNIT IS ESSENTIAL FOR TIGHT-BINDING OF PSI-C

Removal of the peripheral subunits PSI-C, -D and -E from the photosyst em I (PSI) complex of barley requires a urea treatment much harsher th an required to remove the similar subunits from cyanobacterial PSI. Th e resulting PSI barley core was reconstituted by addition of the E. co li expressed subunits PSI-C and -D, and PSI-E isolated from barley. We stern blotting, flash photolysis and NADP(+) photoreduction measuremen ts demonstrated complete and specific removal of the three subunits fr om 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 trun cated barley PSI-D lacking 24 amino acid residues and thus being witho ut the N-terminal extension characteristic for higher plant PSI-D prot eins reconstitutes the PSI core to 50% of the level obtained with inta ct PSI-D as demonstrated by flash photolysis and NADP(+) photoreductio n measurements. Cyanobacterial PSI-D is functionally equivalent to tru ncated barley PSI-D with respect to its activity to reconstitute the P SI 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-terminu s of PSI-D is hypothesized to execute its function through interaction with a plant-specific PSI subunit, possibly PSI-H. An anchoring funct ion 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 NA DP(+) reduction but does not influence electron transfer to iron-sulph ur 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.