Protein rotational mobility in thylakoid membranes of different polypeptide composition in the wild type and mutant strains of Chlamydomonas reinhardtii.
The rotational mobility of thylakoid membrane proteins labeled with a paramagnetic analog of N-ethylmaleimide was investigated by saturation transfer electron spin resonance. In the wild type strain of Chlamydomonas reinhardtii two polypeptides are prominently labeled. They correspond to the 19-kDa subunit of the reaction center I protein and to the 30-kDa subunit of the light harvesting complex. Several polypeptides, most of which are either trypsin or alkaline sensitive, are also labeled. In order to circumvent the lack of specificity during the labeling, we have compared the rotational mobilities of labeled proteins in thylakoid membranes from several mutant strains which lack in photosystem I., ATPase or light harvesting complexes. Comparison of the saturation transfer electron spin resonance spectra obtained with these mutant membranes as well as with trypsin- and alkaline-treated membranes allowed us to characterize the rotational contribution of some of the labeled proteins to the overall protein dynamics observed in the wild type strain. The reaction center I protein undergoes slow rotation as compared to the other labeled proteins. The rotational characteristics of the labeled light harvesting complexes are those of a peptide fragment in the complex which is in rapid motion in unstacked membranes. Stacking of the thylakoid membranes upon Mg2+ addition is accompanied by a marked change in shape of the saturation transfer spectra, and corresponds to the appearance of highly immobilized nitroxides. We interpret these changes as arising mainly from the hindrance upon membrane appression, of the labeled fragment of the light harvesting complexes which protrude at the thylakoid outer surface.
D002696:Chlamydomonas / Q000235:genetics / Q000378:metabolism*; D002736:Chloroplasts / Q000378:metabolism*; D004578:Electron Spin Resonance Spectroscopy / Q000379:methods; D005033:Ethylmaleimide; D006863:Hydrogen-Ion Concentration; D008274:Magnesium / Q000494:pharmacology; D008301:Maleimides; D008560:Membrane Fluidity; D008565:Membrane Proteins / Q000378:metabolism*; D009154:Mutation; D010446:Peptide Fragments / Q000032:analysis; D010455:Peptides / Q000032:analysis*; D013113:Spin Labels
D008301:Maleimides; D008565:Membrane Proteins; D010446:Peptide Fragments; D010455:Peptides; D013113:Spin Labels; C034994:N-methylmaleimide nitroxide; D008274:Magnesium; D005033:Ethylmaleimide