The stable assembly of newly synthesized PsaE into the photosystem I complex occurring via the exchange mechanism is facilitated by electrostatic interactions.
Photosystem I (PSI) is a photochemically active membrane protein complex that functions at the reducing site of the photosynthetic electron-transfer chain as plastocyanin-ferredoxin oxidoreductase. PsaE, a peripheral subunit of the PSI complex, plays an important role in the function of PSI. PsaE is involved in the docking of ferredoxin/flavodoxin to the PSI complex and also participates in the cyclic electron transfer around PSI. The molecular characterization of the assembly of newly synthesized PsaE in the thylakoid membranes or in isolated PSI complexes is the subject of the present study. For this purpose the Mastigocladus laminosus psaE gene was cloned and overexpressed in Escherichia coli, and the resulting PsaE protein was purified to homogeneity by affinity chromatography. The purified PsaE was then introduced into thylakoids isolated from M. laminosus, and the newly introduced PsaE subunit saturates the membrane. The solubilization and separation of the different thylakoid protein complexes indicated that PsaE accumulates specifically in its functional location, the PSI complex. A similar stable assembly was detected when PsaE was introduced into purified PSI complexes, i.e., in the absence of other thylakoid components. This strongly indicates that the information for the stable assembly of PsaE into PSI lies within the polypeptide itself and within other subunits of the PSI complex that interact with it. To determine the nature of these interactions, the assembly reaction was performed in conditions affecting the ionic/osmotic strength. We found that altering the ionic strength significantly affects the capability of PsaE to assemble into isolated thylakoids or PSI complexes, strongly supporting the fact that electrostatic interactions are formed between PsaE and other PSI subunits. Moreover, the data suggest that the formation of electrostatic interactions occurs concomitantly with an exchange step in which newly introduced PsaE replaces the subunit present in situ.
Lushy
Amit
A
Department of Plant Sciences, The Silberman Institute of Life Sciences, The Hebrew University of Jerusalem, Edmond Safra Campus, Givat Ram, Jerusalem 91904, Israel.
Verchovsky
Lilya
L
Nechushtai
Rachel
R
D000458:Cyanobacteria / Q000737:chemistry / Q000378:metabolism; D045322:Photosynthetic Reaction Center Complex Proteins / Q000096:biosynthesis* / Q000737:chemistry* / Q000302:isolation & purification / Q000378:metabolism*; D045331:Photosystem I Protein Complex; D011499:Protein Processing, Post-Translational; D021122:Protein Subunits; D055672:Static Electricity; D020524:Thylakoids / Q000737:chemistry / Q000378:metabolism
D045322:Photosynthetic Reaction Center Complex Proteins; D045331:Photosystem I Protein Complex; D021122:Protein Subunits; C079741:photosystem I, psaE subunit