| 1. |
- Blomqvist, Lisa A., 1978, et al.
(författare)
-
Proteomic analysis of highly purified prolamellar bodies reveals their significance in chloroplast development
- 2008
-
Ingår i: Photosynthesis Research. - : Springer Science and Business Media LLC. - 0166-8595 .- 1573-5079. ; 96:1, s. 37-50
-
Tidskriftsartikel (refereegranskat)abstract
- The prolamellar body (PLB) proteome of dark-grown wheat leaves was characterized. PLBs are formed not only in etioplasts but also in chloroplasts in young developing leaves during the night, yet their function is not fully understood. Highly purified PLBs were prepared from 7-day-old dark-grown leaves and identified by their spectral properties as revealed by low-temperature fluorescence spectroscopy. The PLB preparation had no contamination of extra-plastidal proteins, and only two envelope proteins were found. The PLB proteome was analysed by a combination of 1-D SDS-PAGE and nano-LC FTICR MS. The identification of chlorophyll synthase in the PLB fraction is the first time this enzyme protein was found in extracts of dark-grown plants. This finding is in agreement with its previous localization to PLBs using activity studies. NADPH:protochlorophyllide oxidoreductase A (PORA), which catalyses the reduction of protochlorophyllide to chlorophyllide, dominates the proteome of PLBs. Besides the identification of the PORA protein, the PORB protein was identified for the first time in dark-grown wheat. Altogether 64 unique proteins, representing pigment biosynthesis, photosynthetic light reaction, Calvin cycle proteins, chaperones and protein synthesis, were identified. The in number of proteins' largest group was the one involved in photosynthetic light reactions. This fact strengthens the assumption that the PLB membranes are precursors to the thylakoids and used for the formation of the photosynthetic membranes during greening. The present work is important to enhance our understanding of the significance of PLBs in chloroplast development.
|
|
| 2. |
- Blomqvist, Lisa A., 1978, et al.
(författare)
-
Proteomic analysis of the etioplast inner membranes of wheat (Triticum aestivum) by two-dimensional electrophoresis and mass spectrometry
- 2006
-
Ingår i: Physiologia Plantarum. - : Wiley. - 0031-9317. ; 128:2, s. 368-381
-
Tidskriftsartikel (refereegranskat)abstract
- The proteome of the etioplast inner membranes (EPIM) of dark-grown wheat leaves (Triticum aestivum L.) was mapped as an essential part of studies on plastid differentiation. Proteins were separated by two-dimensional gel electrophoresis and analysed with mass spectrometry (MS). Over 200 protein spots were resolved and visualized by Coomassie blue staining. More than 100 spots were submitted for subsequent mass spectrometry analyses by matrix-assisted laser desorption ionizationtime of flight (MALDI-ToF) MS, electrospray tandem MS (ESI-MS/MS) or liquid chromatographymass spectrometry (LC-MS/MS). There were 46 identified spots, from which at least 21 different proteins were identified. Among these were FtsH proteases and the peptidyl-prolyl cistrans isomerase TLP40, as well as chloroplast coupling factor subunits and extrinsic subunits of photosystem II (PSII). Of special interest is the NADPH:protochlorophyllide oxidoreductase (POR), which is the predominant protein of prolamellar bodies, where it accumulates in a highly stable ternary complex with protochlorophyllide and NADPH. This complex is known to play an important role in the formation and dispersal of prolamellar bodies. Five different isoforms of POR, with different pI values, were identified. We discuss the possibility of these isoforms being differently phosphorylated as part of the regulation of PORpigment complexes. The proteome mapping of EPIM is a crucial step in the understanding of the light-dependent transition of etioplasts to chloroplasts, and provides a basis for functional studies on factors influencing the greening process.
|
|
| 3. |
- Harsanyi, Anett, et al.
(författare)
-
Alterations of NADPH:protochlorophyllide oxidoreductase quantity and lipid composition in etiolated barley seedlings infected by Barley stripe mosaic virus (BSMV)
- 2006
-
Ingår i: Molecular Plant Pathology. - : Wiley. - 1464-6722 .- 1364-3703. ; 7:6, s. 533-541
-
Tidskriftsartikel (refereegranskat)abstract
- To understand the phenomenon by which infection of seed-transmitted Barley stripe mosaic virus (BSMV) alters membrane structures and inhibits protochlorophyllide biosynthesis of dark-grown barley (Hordeum vulgare L.) plants, we analysed the presence of NADPH:protochlorophyllide oxidoreductase (POR, EC 1.3.1.33) and the galactolipid content and fatty acid composition. The amount of POR in etioplasts of infected leaves, compared with non-infected leaves, was reduced, as measured by immunoelectron microscopy and Western blot. These results are in agreement with the previously described reduction of the ratio of the photoactive 650 nm to non-photoactive 630 nm absorbing protochlorophyllide forms (Harsányi et al., 2002. Physiol. Plant 114, 149155). The galactolipid content was lower in infected leaves. Monogalactosyl-diacylglycerol (MGDG) content was reduced to 40% and digalactosyl-diacylglycerol to 55% of control plants on a fresh weight basis. In infected plants, the proportion of linolenic acid decreased in both galactolipids. The lower amount of highly unsaturated fatty acids and the reduced abundance of MGDG correlated well with the previously detected reduction in the membrane ratio of prolamellar body (PLB) to prothylakoid (Harsányi et al., 2002. Physiol. Plant 114, 149155). The reduced amount of POR and the above described alterations in the lipid composition resulted in a disturbed structure of PLBs. As a consequence, pigment synthesis and the greening process were inhibited in infected cells, in turn explaining the appearance of chlorotic stripes of BSMV-infected barley leaves. Our results show that BSMV infection can be detected at a very early stage of leaf development.
|
|
| 4. |
- Solymosi, K., et al.
(författare)
-
Molecular rearrangement in POR macrodomains as a reason for the blue shift of chlorophyllide fluorescence observed after phototransformation
- 2007
-
Ingår i: Biochimica Et Biophysica Acta-Biomembranes. - : Elsevier BV. - 0005-2736. ; 1768:6, s. 1650-1658
-
Tidskriftsartikel (refereegranskat)abstract
- The activation energy and activation volume of the spectral blue shift subsequent to protochlorophyllide phototransformation (called Shibata shift in intact leaves) were studied in prolamellar body (PLB) and prothylakoid-(PT)-enriched membrane fractions prepared from dark-grown wheat (Triticum aestivum, L.) leaves. The measurements were done at 20, 30 and 40 degrees C and at various pressure values. The activation energy values were 181 +/- 8 kJ mol(-1) and 188 +/- 6 kJ mol(-1) for the PLBs and the PTs, respectively. The pressure stabilized the structure of the NADPH: protochlorophyllide oxidoreductase (POR) macrodomains; it prevented or slowed down the blue shift. There were no significant differences between the activation volumes of PLBs and PTs at 30 or 40 degrees C giving values around 100-125 ml mol(-1) which correspond to changes in the tertiary structure of proteins but also resemble the volume changes occurring during the disaggregation of protein dimers or oligomers, or during dissociation of peripheral membrane proteins from membranes. The small differences in the activation parameters of PLBs and PTs indicate that molecular rearrangements inside the POR macrodomains are the primary reasons of the fluorescence blue shift; however, their lipid microenvironment must be also important in the initialization of the shift. (c) 2007 Elsevier B.V. All rights reserved.
|
|
