Photochemical Competence of Assembled Photosystem II Core Complex in Cyanobacterial Plasma Membrane

Michelle Liberton,Himadri B. Pakrasi,Nir Keren

doi:10.1074/jbc.m410218200

Michelle Liberton, Himadri B. Pakrasi + Show 1 more

Open Access

https://doi.org/10.1074/jbc.m410218200

Copy DOI

Abstract

Cyanobacterial cells have two autonomous internal membrane systems, plasma membrane and thylakoid membrane. In these oxygenic photosynthetic organisms the assembly of the large membrane protein complex photosystem II (PSII) is an intricate process that requires the recruitment of numerous protein subunits and cofactors involved in excitation and electron transfer processes. Precise control of this assembly process is necessary because electron transfer reactions in partially assembled PSII can lead to oxidative damage and degradation of the protein complex. In this communication we demonstrate that the activation of PSII electron transfer reactions in the cyanobacterium Synechocystis sp. PCC 6803 takes place sequentially. In this organism partially assembled PSII complexes can be detected in the plasma membrane. We have determined that such PSII complexes can undergo light-induced charge separation and contain a functional electron acceptor side but not an assembled donor side. In contrast, PSII complexes in thylakoid membrane are fully assembled and capable of multiple turnovers. We conclude that PSII reaction center cores assembled in the plasma membrane are photochemically competent and can catalyze single turnovers. We propose that upon transfer of such PSII core complexes to the thylakoid membrane, additional proteins are incorporated followed by binding and activation of various donor side cofactors. Such a stepwise process protects cyanobacterial cells from potentially harmful consequences of performing water oxidation in a partially assembled PSII complex before it reaches its final destination in the thylakoid membrane.

Highlights

Photosystem II (PSII)1 is an integral membrane chlorophyllprotein complex that is a part of the photosynthetic electron transfer chain in the thylakoid membranes of cyanobacteria and plant chloroplasts
Because active photosynthesis occurs in the thylakoid membranes (TM) we have proposed that the core subunits D1, D2, and cytochrome b559 are assembled in the plasma membranes (PM), and the CP43 and CP47 subunits are added in the TM [8]
During this work we studied the activity of photosystem II (PSII) complexes in PM and TM to reconstruct the pathway of cofactor assembly and activation

Summary

EXPERIMENTAL PROCEDURES

Bacterial Cell Culture Conditions—Synechocystis 6803 cells were grown in BG11 medium [9] at 30 °C under 50 ␮mol of photons mϪ2 sϪ1 white light. Membrane Isolation and Two-phase Partitioning—Frozen cells were thawed on ice and resuspended in a low salt buffer (20 mM potassium phosphate, pH 7.8). Measurements were performed on an equal chlorophyll basis (16 ␮g of Chl/ml). Manganese concentrations in isolated membranes digested in 5% nitric acid were measured using an AA600 atomic absorption spectrometer (PerkinElmer Life Sciences). 1-ml samples containing 25 ␮g of Chl were incubated with radioactive DCMU in the presence or absence of a 0.1 mM excess cold DCMU. The concentration of the radioactive DCMU in the medium was measured by scintillation counting as described in Lind et al [12]. The ratio of the radioactivity measured in the presence and absence of cold DCMU was used to calculate the concentrations of bound and free [14C]DCMU

RESULTS

TABLE I Properties of thylakoid and plasma membranes

The water splitting activity on the donor side of PSII relies

DISCUSSION