| 1. |
- Arvidsson, Per-Ola, et al.
(författare)
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Violaxanthin accessibility and temperature dependency for de-epoxidation in spinach thylakoid membranes
- 1997
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Ingår i: Photosynthesis Research. - 0166-8595. ; 52:1, s. 39-48
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Tidskriftsartikel (refereegranskat)abstract
- Using DTT and iodoacetamide as a novel irreversible method to inhibit endogenous violaxanthin de-epoxidase, we found that violaxanthin could be converted into zeaxanthin from both sides of the thylakoid membrane provided that purified violaxanthin de-epoxidase was added. The maximum conversion was the same from both sides of the membrane. Temperature was found to have a strong influence both on the rate and degree of maximal violaxanthin to zeaxanthin conversion. Thus only 50% conversion of violaxanthin was detected at 4 degreesC, whereas at 25 degreesC and 37 degreesC the degree of conversion was 70% and 80%, respectively. These results were obtained with isolated thylakoids from non-cold acclimated leafs. Pigment analysis of sub-thylakoid membrane domains showed that violaxanthin was evenly distributed between stroma lamellae and grana partitions. This was in contrast to chlorophyll a and beta-carotene which were enriched in stroma lamellae fractions while chlorophyll b, lutein and neoxanthin were enriched in the grana membranes. In combination with added violaxanthin de-epoxidase we found almost the same degree of conversion of violaxanthin to zeaxanthin (73-78%) for different domains of the thylakoid membrane. We conclude that violaxanthin de-epoxidase converts violaxanthin in the lipid matrix and not at the proteins, that violaxanthin does not prefer one particular membrane region or one particular chlorophyll protein complex, and that the xanthophyll cycle pigments are oriented in a vertical manner in order to be accessible from both sides of the membrane when located in the lipid matrix.
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| 2. |
- Jansson, Stefan, et al.
(författare)
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Antenna protein composition of PS I and PS II in thylakoid sub-domains
- 1997
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Ingår i: Biochimica et Biophysica Acta - Bioenergetics. - 0005-2728. ; 1320:3, s. 297-309
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Tidskriftsartikel (refereegranskat)abstract
- Spinach thylakoids were separated into grana core, grana margin, and two different stromal lamella fractions in the absence of detergents. The levels of all light-harvesting chlorophyll a/b-binding (LHC) proteins were determined in all fractions, and were normalised to the amount of Photosystem I (PS I) and Photosystem II (PS II) centres. PS Iβ in the stroma lamellae was found to have a full complement of Lhca polypeptides and, probably, one attached LHC II trimer. PS Iα binds additional LHC II trimers, but PS I centres located in the inner parts of the grana stack lack Lhca1 and are depleted in Lhca4. PS IIβ, found in grana margins and stroma lamellae, seems to associate one monomer each of Lhcb4, Lhcb5 and Lhcb6 (CP29, CP26 and CP24, respectively) and one LHC II trimer consisting of two Lhcb1 and one Lhcb3 subunit. PS IIα has additional LHC II trimers (consisting of Lhcb1 and Lhcb2) attached. We also find evidence for the existence of both PS I and PS II centres in the extreme stroma (probably centres being synthesised or repaired), that lack all LHC proteins.
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| 3. |
- Mamedov, Fikret, et al.
(författare)
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Photosystem II in Different Parts of the Thylakoid Membrane: A Functional Comparison between Different Domains
- 2000
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Ingår i: Biochemistry. - : American Chemical Society (ACS). - 0006-2960 .- 1520-4995. ; 39:34, s. 10478-10486
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Tidskriftsartikel (refereegranskat)abstract
- The electron transport properties of photosystem II (PSII) from five different domains of the thylakoid membrane were analyzed by flash-induced fluorescence kinetics. These domains are the entire grana, the grana core, the margins from the grana, the stroma lamellae, and the Y100 fraction (which represent more purified stroma lamellae). The two first fractions originate from appressed grana membranes and have PSII with a high proportion of O2-evolving centers (80-90%) and efficient electron transport on the acceptor side. About 30% of the granal PSII centers were found in the margin fraction. Two-thirds of those PSII centers evolve O2, but the electron transfer on the acceptor side is slowed. PSII from the stroma lamellae was less active. The fraction containing the entire stroma has only 43% O2-evolving PSII centers and slow electron transfer on the acceptor side. In contrast, PSII centers of the Y100 fraction show no O2 evolution and were unable to reduce QB. Flash-induced fluorescence decay measurements in the presence of DCMU give information about the integrity of the donor side of PSII. We were able to distinguish between PSII centers with a functional Mn cluster and without any Mn cluster, and PSII centers which undergo photoactivation and have a partially assembled Mn cluster. From this analysis, we propose the existence of a PSII activity gradient in the thylakoid membrane. The gradient is directed from the stroma lamellae, where the Mn cluster is absent or inactive, via the margins where photoactivation accelerates, to the grana core domain where PSII is fully photoactivated. The photoactivation process correlates to the PSII diffusion along the membrane and is initiated in the stroma lamellae while the final steps take place in the appressed regions of the grana core. The margin domain is seemingly very important in this process.
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| 4. |
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| 5. |
- Stefánsson, Hreinn, et al.
(författare)
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Fractionation of the thylakoid membrane from Dunaliella salina – heterogeneity is found in Photosystem I over a broad range of growth irradiance
- 1997
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Ingår i: Biochimica et Biophysica Acta - Bioenergetics. - 0005-2728. ; 1320:3, s. 235-246
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Tidskriftsartikel (refereegranskat)abstract
- Thylakoids from the green alga, Dunaliella salina, were fragmented by sonication and the appressed grasna membranes separated from stroma lamellae by partitioning in aqueous two-phase systems. The concentration and antenna size of Photosystem I in the two membrane domains were determined for cultures grown at three different light intensities. Although the antenna size of both PS is decreased with increasing growth irradiance, the antenna size of Photosystem I in the grana was approximately 25-30% greater than the antenna size of Photosystem I in the stroma lamellae. Counter-current distribution analysis of sonicated thylakoids revealed that the amount of stroma lamellae increased whereas the amount of the stacked membranes decreased at higher growth irradiance. The overall decrease in the antenna size of PS I, in D. salina, at higher light intensities can therefore be explained by the combined effect of a decrease in the antenna size of Photosystem I, both in the grana and the stroma lamellae, and a relative increase in the amount of stroma lamellae which has smaller Photosystem I antennae than the Photosystem I centers found in the grana. Light-induced protein phosphorylation increased the relative amount of the stroma lamella fraction. This is interpreted as a result of partial unstacking of the grana. It is suggested that this may be a mechanism for increasing the cyclic electron transport around Photosystem I.
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| 6. |
- Stefánsson, Hreinn, et al.
(författare)
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Phosphorylation of thylakoids and isolated subthylakoid vesicles derived from different structural domains of the thylakoid membrane from spinach chloroplast
- 1995
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Ingår i: Biochimica et Biophysica Acta - Bioenergetics. - 0005-2728. ; 1231:3, s. 323-334
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Tidskriftsartikel (refereegranskat)abstract
- (1) Thylakoids from spinach chloroplasts were phosphorylated, fragmented by sonication, and then fractionated by aqueous two-phase partitioning to yield membrane fragments, deriving from different structural domains of the membrane: grana, grana margins, grana core and stroma lamellae. The photochemical activities of PS I and PS II, located in the grana, and PS I and PS II, located in the stroma lamellae, were compared for phosphorylated and control thylakoids. The antenna size (reflected by the Km value) and maximum activity (Vmax) of PS II declined by 19 and 23%, respectively, while for PS II the antenna size and Vmax decreased by 4 and 12%, respectively. No significant changes in antenna size were detected for either grana PS I or stroma lamellae PS I. Counter-current distribution was used for the quantitative separation of grana and stroma lamellae vesicles. Upon phosphorylation, the stroma lamellae fraction increased from 30% to 35% of the total, based on total absorbance at 680 nm. This increase can be explained by partial unstacking of the grana periphery and appressed membranes near the fret regions. Portions of the previously stacked membranes can therefore break and separate with the stroma exposed membrane. In addition, since the grana margins contain PS I (with 40% larger antennae than PS I), which is functionally connected to LHC II, it is to be expected that some of these PS I units will also enter the stroma lamellae fraction and thus help contribute to a lower chlorophyll a/b ratio and a small increase in the average PS I antenna size of the stroma lamellae fraction from phosphorylated thylakoids. It is concluded that the incidence of partial destacking of the grana, which occurs due to the phosphorylation of LHC II and PS II polypeptides, may promote the exposure of the granal PS I centers to the aqueous stroma and increase cyclic electron flow around Photosystem I and thereby ATP production over NADPH production. (2) Subthylakoid vesicles, representing the different structural domains, were also examined for their properties following an incubation in presence of light and ATP. Phosphorylation of membrane proteins including LHC II and PS II associated polypeptides was observed in membrane fractions deriving from the grana lamellae and, to a lesser extent, the grana core. Three unidentified polypeptides of 15, 20 and 22 kDa were the most abundantly labeled polypeptides in the stroma lamellae fraction. No membrane proteins became phospho-labeled in the grana margin fraction.
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