Structural Organization of the Major Subunits in Cyanobacterial Photosystem 1

Abstract

Based on an improved isolation procedure using perfusion chromatography, trimeric Photosystem 1 (PS1) complexes have been isolated from various deletion mutants of the mesophilic cyanobacterium Synechocystis PCC 6803. These mutants are only deficient in the deleted subunits, which was carefully checked by high resolution gel electrophoresis in combination with immunoblotting. These highly purified and well characterized PS1 particles were then examined by electron microscopy, followed by computer-aided image processing with single particle averaging techniques as described earlier (Kruip, J., Boekema, E. J., Bald, D., Boonstra, A. Photosystem 1 (PS1) 1 is one of the two transmembrane pigment-protein complexes of oxygenic photosynthesis located in the thylakoid membrane of chloroplasts and cyanobacteria. Its main function is the light-dependent electron transfer from plastocyanin (or cytochrome c 6 ) on the lumen side to soluble ferredoxin (or flavodoxin) on the cytoplasmic or stroma side, thereby transforming light energy into reducing power for various biosynthetic pathways (see Refs. 1-3 for reviews). Cyanobacterial PS1 consists of 11 subunits, named PsaA to PsaM, with PsaG and PsaH missing, as they are found only in chloroplasts (3); all their sequences are known (1). The hydrophilic subunits PsaC, -D, and -E are localized on the cytoplasmic side (4), and PsaF is supposed to sit on the lumenal side of the thylakoid membrane. All the other subunits are integral membrane proteins.In terms of function the initial steps of light harvesting and charge separation are carried out by only three proteins. The PsaA/PsaB heterodimer harbors the primary electron donor P700 (a chlorophyll dimer) and the electron acceptors A 0 (a monomeric chlorophyll), A 1 (a phylloquinone), and F X (a [4Fe4S] iron-sulfur cluster) in addition to about 100 -130 antenna chlorophyll molecules (5). PsaC contains the terminal electron acceptors F A and F B (both [4Fe4S] iron-sulfur clusters). The other proteins do not bind any cofactors, but fulfill important functional and structural roles: PsaD and -F have been shown to be candidates for ferredoxin (6, 7) and plastocyanin docking (8, 9), respectively. PsaE plays an important role in cyclic electron flow (10) and ferredoxin reduction (11,12), while PsaL and PsaD are important for trimer formation (4,13,14).To elucidate the function of all subunits on a molecular level, a high resolution structure is necessary. Despite several crystallization reports (15-17), a low-resolution x-ray analysis (6 Å) for PS1 from the thermophilic cyanobacterium Synechococcus elongatus became available only in 1993 (18). Recently this x-ray analysis was extended to 4 Å (19, 57). In this study, 31 transmembrane, 9 surface, and 3 stromal ␣-helices, 65 chlorophyll a molecules, and three [4Fe4S] clusters could be fitted into the electron density map. Due to the medium resolution it is not possible to attribute the identified structural elements * The costs of publication of this article were defrayed in part by the pay...

Untitled

“…PS I monomers and trimers from Gloeobacter violaceus closely resemble previously analysed PS I complexes from other cyanobacteria like Synechocystis PCC 6803 and Synechococcus elongatus regarding size and shape (Rögner et al 1990;Kruip et al 1997). However, the T-shaped complex in the preparation was never observed in any of the previously studied PS I samples (Kruip et al 1997).…”

“…However, the T-shaped complex in the preparation was never observed in any of the previously studied PS I samples (Kruip et al 1997). Its shape is distantly reminiscent of a recently described TolC protein from gram-negative bacteria spanning both the outer membrane and the periplasmic space (Koronakis et al 2000).…”

“…The elution profile was recorded at 280 and 435 nm. For calibration of the Superose 6 column pure preparations of trimeric and monomeric PS I from Synechocystis or Spirulina platensis were used (Kruip et al 1997(Kruip et al , 1999.…”

“…electron carrier ferredoxin (or alternatively to flavodoxin in cyanobacteria) and ultimately to NADP + to generate NADPH (Chitnis 1996). Typically, cyanobacterial PS I shows a highly conserved subunit pattern consisting of 11 different polypeptides named PsaA-PsaF and PsaI-PsaM (Kruip et al 1997). In cyanobacteria, PS I can occur in two different oligomeric states, a monomeric and a trimeric form (Boekema et al 1987), which seem to be at least partly interchangeable, with a majority of trimers being present in vivo under moderate light conditions (Kruip et al 1994(Kruip et al , 1999).…”

“…In cyanobacteria, PS I can occur in two different oligomeric states, a monomeric and a trimeric form (Boekema et al 1987), which seem to be at least partly interchangeable, with a majority of trimers being present in vivo under moderate light conditions (Kruip et al 1994(Kruip et al , 1999). Trimerisation appears to be dependent on the electrochemical situation in the lipid bilayer and to require the presence of the stroma-exposed subunit PsaD and the membrane integral subunit PsaL (Kruip et al 1999;Chitnis and Chitnis 1997;. PS I complexes from a whole range of cyanobacterial strains have been studied extensively in the past years.…”