Retraction: A proteomic approach for investigation of photosynthetic apparatus in plants

Abstract

The proteome of the photosynthetic apparatus of barley (Hordeum vulgare), obtained by analysis of thylakoids without any previous fractionation, was mapped by native electrophoresis followed by sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) as the second dimension two-dimensional-blue native (2-D/BN)/SDS-PAGE). This protocol provided an excellent alternative to the 2-D-isoelectric focusing/sodium dodecyl sulfate-polyacrylamide gel electrophoresis for 2-D separation of the most hydrophobic thylakoid proteins. Monocots and dicots showed significant differences in the first dimension while in the second dimension patterns appeared similar. Identification of each spot was performed by internal peptide primary sequence determination using both nano-electrospray ionization tandem mass spectrometry and, to a lesser extent, peptide mass fingerprinting matrix-assisted laser desorption/ionization-time of flight using MALDI-TOF. This is due in particular to the fact that a limited number of peptides was obtained after trypsin digestion of these highly hydrophobic proteins. A larger number of peptides from hydrophilic intermembrane domains of transmembrane proteins were detected. Despite this, about 70% of the expected proteins were identified, including proteins with grand average of hydropathicity scores higher than 0.5. It is therefore reasonable to assert that protein hydrophobicity is not the limiting factor. Small proteins were not well identified with trypsin digestion. Instead some of these could be identified using acid hydrolysis. The method presented here does not require prefractionation of different thylakoid complexes and consequently gives confidence in comparing the proteome of the photosynthetic apparatus before and after treatment. It thus allows us to understand the molecular mechanisms underlying physiological adaptations of higher plants and to perform screening of photosynthetic mutants.

l-Galactono-1,4-lactone Dehydrogenase Is Required for the Accumulation of Plant Respiratory Complex I

“…In one-dimensional BN-profiles of leaves, main bands represented complexes originating from thylakoid membranes, which were identifiable from their constitutive spots appearing in the second dimension (Fig. 4, A and B), in agreement with the previous reports of proteomic investigation of plant photosystems (34,35). The presence of mt CI was clearly revealed by in-gel NADH/ NBT staining of the BN profiles from dodecyl maltoside-solubilized proteins of Col-0 leaf membrane extracts (Fig.…”

Structural Organization of Photosynthetic Apparatus in Agranal Chloroplasts of Maize

“…1). As expected, our results of BN-and BN/SDS-PAGE confirm that the protein composition of the C 4 MS membranes is similar to that of the C 3 thylakoids, where all of the complexes listed above are present (24,30,35,36). Our BN/SDS-PAGE identification of the LHCII⅐PSII supercomplexes in the BS agranal membranes (Fig.…”

“…Protein profiles obtained from various plant species using two-dimensional BN/SDS-PAGE are similar due to the high conservation of the subunits of the photosynthetic complexes (24,30,35,36). We therefore identified the components of the BN bands from MS and BS thylakoids by comparison of the apparent molecular weight on BN/SDS-PAGE and by immunodetection using specific antibodies against the subunits of the PSII and PSI cores and the associated LHCII and LHCI antennae (see Fig.…”

Oleosin Is Bifunctional Enzyme That Has Both Monoacylglycerol Acyltransferase and Phospholipase Activities

“…In this study, we purified the microsomal membranebound MGAT from immature peanut seeds using a combination of urea (a chaotropic agent) and CHAPS (a zwitterionic detergent) without the loss of enzyme activity. A simple and functionally active multiprotein complex isolation procedure was developed using native-PAGE (31).…”