Identification and Characterization of an Assembly Intermediate Subcomplex of Photosystem I in the Green Alga Chlamydomonas reinhardtii

Abstract

Photosystem I (PSI) is a multiprotein complex consisting of the PSI core and peripheral light-harvesting complex I (LHCI) that together form the PSI-LHCI supercomplex in algae and higher plants. The supercomplex is synthesized in steps during which 12-15 core and 4 -9 LHCI subunits are assembled. Here we report the isolation of a PSI subcomplex that separated on a sucrose density gradient from the thylakoid membranes isolated from logarithmic growth phase cells of the green alga Chlamydomonas reinhardtii. Pulse-chase labeling of total cellular proteins revealed that the subcomplex was synthesized de novo within 1 min and was converted to the mature PSI-LHCI during the 2-h chase period, indicating that the subcomplex was an assembly intermediate. The subcomplex was functional; it photooxidized P700 and demonstrated electron transfer activity. The subcomplex lacked PsaK and PsaG, however, and it bound PsaF and PsaJ weakly and was not associated with LHCI. It seemed likely that LHCI had been integrated into the subcomplex unstably and was dissociated during solubilization and/or fractionation. We, thus, infer that PsaK and PsaG stabilize the association between PSI core and LHCI complexes and that PsaK and PsaG bind to the PSI core complex after the integration of LHCI in one of the last steps of PSI complex assembly.

Biogenic membranes of the chloroplast in Chlamydomonas reinhardtii

“…3 F and G). This result and previously reported evidence that this complex is a late intermediate in PSI assembly (21), suggest that later steps in PSI assembly occur primarily in thylakoid membranes. Earlier steps in PSI assembly may also occur in thylakoids because we detected an early PSI assembly factor, YCF4, only in thylakoid membrane fractions (Fig.…”

“…3 F and G). Although PsaAp was not detected unassembled, it was detected in the PSI monomeric complex, the PSI monomeric complex lacking PsaK and PsaG, and a larger unidentified complex of approximately 550 kDa, possibly the PSI dimer (21). Notably, the PSI monomer lacking PsaK/G was more abundant in the thylakoid membrane fraction than in the CTM fraction (Fig.…”

Regulatory factors for the assembly of thylakoid membrane protein complexes

“…Following the binding of co-factors to the two reaction centre proteins, the PsaA/B reaction centre heterodimer is formed [79]. Subsequently, relatively small subunits with a maximum molecular mass of 18 kDa are attached to the heterodimer [80].…”

Regulation of the photosynthetic apparatus under fluctuating growth light

“…PsaC, which itself contains two Fe-S clusters that are incorporated in the stroma, can only assemble with a PsaA/B heterodimer that already contains the Fx Fe-S cluster [53,54]. Next, the assembly of the other stromal subunits PsaD and PsaE takes place, followed by the assembly of the remaining subunits and finally the LHCI antenna [50,51]. As with PSII, several auxiliary proteins involved in PSI assembly have been identified [50].…”

“…One obstacle in research into PSI assembly in plant chloroplasts has been the fast rate at which this occurs [50,51]. Nevertheless, it is known from Chlamydomonas that the biogenesis of PSI is controlled by the epistasy of synthesis process (CES), in which the presence of the PsaB protein enables the synthesis of the PsaA protein, which is in turn required for translation of the PsaC protein [52].…”