The kinetics of P515 in relation to the lipid composition of the thylakoid membrane

Flash-induced P515 absorbance changes have been studied in dark-adapted chloroplasts isolated from spinach plants grown under two different light intensities. The slow component (reaction 2), normally present in the P515 response of chloroplasts isolated from plants grown at an intensity of 60 W · m^–2, was largely reduced in chloroplasts isolated from plants grown at an intensity of 6 W · m^–2. This reduction of the slow component in the P515 response appeared to be coincident with an alteration in the lipid composition of the thylakoid membrane. Mainly the ratio monogalactosyldiacylglycerol to digalactosyldiacylglycerol appeared to be altered. In thylakoids from plants grown at 6 W · m^–2, the ratio was approximately 35% lower than that of plants grown at 60 W · m^–2. The amount of both cytochromeb ₅₆₃ and cytochromef was largely reduced in chloroplasts isolated from plants grown at low light intensity. These results may indicate a possible correlation between structural organization of the thylakoid membrane and the kinetics of the flash-induced P515 response.     

Development of Oat Prothylakoids into Thylakoids during Greening Does Not Change Transmembrane Galactolipid Asymmetry but Preserves the Thylakoid Bilayer

The lipase from Rhizopus arrhizus and the lipolytic acyl hydrolase from potato tubers have been used to determine the transmembrane distribution of monogalactosyldiacylglycerol (MGDG) and digalactosyldiacylglycerol (DGDG) in prothylakoids and thylakoids from oat (Avena sativa). Both galactolipids were found to be asymmetrically distributed. The molar outside/inside distribution was 70 ± 8/30 ± 8 for MGDG and 10 ± 4/90 ± 4 for DGDG in the prothylakoid membrane. Mature thylakoids presented a similar distribution, i.e. 63 ± 4/37 ± 4 for MGDG and 12 ± 3/88 ± 3 for DGDG. This distribution has been assessed under a variety of different conditions, namely (a) in media favoring thylakoid stacking or unstacking and inducing various membrane surface potentials, (b) in the presence of defatted bovine serum albumin which removed free fatty acids and partially lyso-galactolipids, (c) under various temperature conditions which resulted in different hydrolysis rates and degrees of fluidity of the membrane, and (d) in the presence of different enzyme concentrations which influenced the hydrolysis rate. The above distribution was found to be independent of the type of conditions used. Nonbilayer forming/bilayer forming lipid ratios suggest that both monolayers of the prothylakoid and the inner monolayer of oat thylakoid membranes should display lamellar structures (e.g. ratios <2.5). In contrast the outer monolayer of the thylakoid membrane should display non-lamellar configurations (e.g. ratio >2.5). Thus, it is concluded that the incorporation of chlorophyll-protein complexes into the nascent thylakoid membrane modifies neither the galactolipid nor the phospholipid transmembrane distribution. However, these complexes appear to be crucial to preserve a bilayer configuration to the greening membrane which, otherwise, would adopt nonlamellar structures. The possible origin of galactolipid transversal asymmetry which appears very early during the biogenesis of oat thylakoid membranes is discussed.     

Characteristics of chloroplast thylakoid lipid composition associated with resistance to triazine herbicides

A detailed comparison of the polar-lipid composition of chloroplast thylakoid membranes isolated from triazine-susceptible and triazine-resistant biotypes of Chenopodium album, Senecio vulgaris, Poa annua and Amaranthus retroflexus has been carried out. No major differences in the composition of the bulk lipid matrix were found except for a slightly higher monogalactosyldiacylglycerol to digalactosyldiacylglycerol ratio in resistant compared with susceptible biotypes. There was, however, in the case of resistant plants a higher level of phosphatidylglycerol-containing transhexadecenoic acid in membrane fractions enriched in photosystem two. It is concluded that although the minor differences could contribute to triazine resistance it is more likely that they reflect secondary alterations in membrane organisation associated with changes in relative levels of pigment-protein complexes.Abbreviations DGDG digalactosyldiacylglycerol - MGDG monogalactosyldiacylglycerol - PG phosphatidylglycerol - PSII photosystem two     

Human complement subcomponent C2: purification and proteolytic cleavage in fluid phase by C1s, C1r2-C1s2 and C1

Photosynthetic membranes contain considerable regions of high surface curvature, notably at their margins, where the average radius of curvature is about 10 nm. The proportion of total membrane lipid in the outer and inner thylakoid margin monolayers is estimated at 21% and 13%, respectively. The major thylakoid lipid, monogalactosyldiacylglycerol, is roughly cone-shaped and will not form complete lamellar bilayer phases, even in combination with other thylakoid lipids. It is proposed that this galactolipid plays a role in: (a) stabilising regions of concave curvature in thylakoids; and (b) packaging hydrophobic proteins in planar bilayer regions by means of inverted micelles. This model predicts substantial asymmetries in the distribution of lipids both across and along the thylakoid bilayer plane.     

The effect of low oxygen concentration on the acyl-lipid and fatty-acid composition of the C4 plant Amaranthus paniculatus L.

Acyl lipids and their constituent fatty acids were studied in leaves, chloroplasts and bundle-sheath strands of the C₄ plant Amaranthus paniculatus L. grown under normal and 4%-oxygen-containing atmospheres. In all fractions the major lipids were found to be monogalactosyldiacylglycerol, digalactosyldiacylglycerol, sulphoquinovo-syldiacylglycerol and phosphatidylglycerol. Significant quantities of phosphatidylcholine and phosphatidylethanolamine were restricted to leaves and bundle-sheath strands. All lipids, except phosphatidylglycerol where 3-trans-hexadecenoic acid was also present, contained palmitic acid, stearic acid, oleic acid, linoleic acid and linolenic acid. On a chlorophyll basis and compared with whole leaves, the amounts of phosphatidylcholine and phosphatidylethanolamine in bundle-sheath strands were considerably reduced. Three weeks after the change from a normal to a 4% atmospheric O₂ level, the galactolipid content, particularly in the bundlesheath strands, was enhanced. There were no significant differences in the degrees of saturationunsaturation of total acyl lipid for the plants grown in the low oxygen and normal atmospheres, although under 4% O₂ the phosphatidylglycerol contained an increased proportion of 3-trans-hexadecenoic acid at the expense of palmitic acid.Abbreviations DGDG digalactosyldiacylglycerol - MGDG monogalactosyldiacylglycerol - PC phosphatidylcholine - PE phosphatidylethanolamine - PG phosphatidylglycerol - SQDG sulphquinovosyldiacylglycerol     

Chloroplast biogenesis. Regulation of lipid transport to the thylakoid in chloroplasts isolated from expanding and fully expanded leaves of pea

To study the regulation of lipid transport from the chloroplast envelope to the thylakoid, intact chloroplasts, isolated from fully expanded or still-expanding pea (Pisum sativum) leaves, were incubated with radiolabeled lipid precursors and thylakoid membranes subsequently were isolated. Incubation with UDP[(3)H]Gal labeled monogalactosyldiacylglycerol in both envelope membranes and digalactosyldiacylglycerol in the outer chloroplast envelope. Galactolipid synthesis increased with incubation temperature. Transport to the thylakoid was slow below 12 degrees C, and exhibited a temperature dependency closely resembling that for the previously reported appearance and disappearance of vesicles in the stroma (D.J. Morré, G. Selldén, C. Sundqvist, A.S. Sandelius [1991] Plant Physiol 97: 1558-1564). In mature chloroplasts, monogalactosyldiacylglycerol transport to the thylakoid was up to three times higher than digalactosyldiacylglycerol transport, whereas the difference was markedly lower in developing chloroplasts. Incubation of chloroplasts with [(14)C]acyl-coenzyme A labeled phosphatidylcholine (PC) and free fatty acids in the inner envelope membrane and phosphatidylglycerol at the chloroplast surface. PC and phosphatidylglycerol were preferentially transported to the thylakoid. Analysis of lipid composition revealed that the thylakoid contained approximately 20% of the chloroplast PC. Our results demonstrate that lipids synthesized at the chloroplast surface as well as in the inner envelope membrane are transported to the thylakoid and that lipid sorting is involved in the process. Furthermore, the results also indicate that more than one pathway exists for galactolipid transfer from the chloroplast envelope to the thylakoid.     

Changes in the binding and inhibitory properties of urea/triazine-type herbicides upon phospholipid and galactolipid depletion in the outer monolayer of thylakoid membranes

The binding characteristics and the inhibitory power of atrazine and DCMU towards uncoupled electron flow activity were studied in acyl lipid-depleted thylakoid membranes from atrazine-susceptible and-resistant biotypes of Solanum nigrum L. For this purpose, phospholipase A₂ from Vipera russelli and the lipase from Rhizopus arrhizus were used to obtain a selective lipid class (phospholipids or galactolipids) depletion which was restricted to the outer monolayer. Neither phospholipid nor galactolipid removal affected the dissociation constant and the number of binding sites of atrazine. In contrast, the dissociation constant of DCMU was increased in phospholipid-depleted thylakoid membranes but remained unchanged after galactolipid depletion. The number of DCMU binding sites decreased significantly after both lipase treatments, but only in the resistant biotype. The inhibitory effectiveness of the herbicide was either decreased or increased (to different extents) depending on the lipid class which was removed from the membrane and on the biotype considered. These results are discussed with reference to the possible conformational changes of the 32 kDa herbicide-binding polypeptide occurring after lipase treatments.Abbreviations Atrazine 2-chloro-4-(ethylamino)-6-(isopropylamino)-s-triazine - BSA bovine serum albumin - DCMU diuron, 3-(3,4-dichlorophenyl)-1,1-dimethylurea - DGDG digalactosyldiacylglycerol - LRa lipase from Rhizopus arrhizus - MGDG monogalactosyldiacylglycerol - PC phosphatidylcholine - PG phosphatidylglycerol - PLA₂ phospholipase A₂ - R atrazine-resistant - S atrazinesusceptible     

Growth temperature effects on thylakoid membrane lipid and protein content of pea chloroplasts

The lipid composition and level of unsaturation of fatty acids has been determined for chloroplast thylakoid membranes isolated from Pisum sativum grown under cold (4°/7°C) or warm (14°/17°C) conditions. Both the relative amounts of lipid classes and degree of saturation were not greatly changed for the two growth conditions. In cold-grown plants, there was a slightly higher linolenic and lower linoleic acid content for the glycolipids monogalactosyldiacylglycerol (MGDG), digalactosyldiacylglycerol (DGDG), and sulfoquinovosyldiacylglycerol. In contrast to thylakoid membranes, a non-thylakoid leaf membrane fraction including the chloroplast envelope, had a higher overall level of fatty acid unsaturation in cold-grown plants due mainly to an increase in the linolenic acid content of MGDG, DGDG, phosphatidylglycerol, and phosphatidylcholine. The most clear cut change in the thylakoid membrane composition was the lipid to protein ratio which was higher in the cold-grown plants.     

Membrane curvature stress controls the maximal conversion of violaxanthin to zeaxanthin in the violaxanthin cycle—influence of α-tocopherol, cetylethers, linolenic acid, and temperature

Zeaxanthin, an important component in protection against overexcitation in higher plants, is formed from violaxanthin by the enzyme violaxanthin de-epoxidase. We have investigated factors that may control the maximal degree of conversion in the violaxanthin cycle. The conversion of violaxanthin to zeaxanthin in isolated spinach thylakoids was followed at different temperatures and in the presence of lipid packing modifiers. The maximum degree of conversion was found to be 35%, 70% and 80% at 4 °C, 25 °C and 37 °C respectively. In the presence of membrane modifying agents, known to promote non-lamellar structures (H_II), such as linolenic acid the conversion increased, and the maximal level of violaxanthin de-epoxidation obtained was close to 100%. In contrast, substances promoting lamellar phases (L_α), such as α-tocopherol and 8-cetylether (C₁₆EO₈), only 55% and 35% of the violaxanthin was converted at 25 °C, respectively. The results are interpreted in light of the lipid composition of the thylakoid membrane, and we propose a model where a negative curvature elastic stress in the thylakoid lipid bilayer is required for violaxanthin de-epoxidase activity. In this model zeaxanthin with its longer hydrophobic stretch is proposed to promote lamellar arrangements of the membrane. As a result, zeaxanthin relieves the curvature elastic stress, which in turn leads to inactivation of violaxanthin de-epoxidase.     

Transverse heterogeneity in lipid fluidity in spinach thylakoids,photosystem II preparations,and thylakoid galactolipid vesicles

We have used three doxyl stearic acid spin labels to study the transverse hetero-geneity in lipid fluidity in thylakoids, photosystem II (PS II) preparations, and thylakoid galactolipid vesicles. This comparative study shows that spin labels incorporated into the membrane of the PS II preparation experience far more immobilization than do the same spin labels incorporated into either thylakoids or vesicles prepared from the polar lipids extracted from thylakoids. The spin label immobilization found in the PS II preparation is manifest even near the center of the bilayer, where lipid mobility is normally at its maximum. Analysis of the lipid content of the PS II preparation, relative to chlorophyll, suggests that the PS II preparation may be lipid depleted. This lipid depletion could explain the results presented. However, electron microscopy [Dunahay et al. (1984) Biochim. Biophys. Acta 764:179–193] has not indicated that major delipidation has occurred, and so it remains possible that the immobilization found in the PS II preparation is due primarily to the normal (but close) juxtaposition of adjacent PS II complexes and the cooperative immobilization of their surrounding lipids. Based on the results presented, we conclude that highly mobile lipids are not required for oxygen evolution, the primary photochemistry or the secondary reduction of exogenously added quinones. Unfortunately, the relationship between the plastoquinone pool and the fluidity of the membrane in the PS II preparation remains ambiguous.Abbreviations PS II photosystem II - SDSA 5-doxylstearic acid - 12DSA 12-doxylstearic acid - 16DSA 16-doxylstearic acid - 7N14 2-heptyl-2-hexyl-5,5-dimethyloxazolidine-N-oxyl - chromium oxalate potassium trioxalatochromiate - EPR electron paramagnetic resonance - Chl chlorophyll - MGDG monogalactosyldiacylglycerol - DGDG digalactosyldiacylglycerol     

Brij detergents reveal new aspects of membrane microdomain in erythrocytes

Membrane microdomains enriched in cholesterol, sphingolipids (rafts), and specific proteins are involved in important physiological functions. However their structure, size and stability are still controversial. Given that detergent-resistant membranes (DRMs) are in the liquid-ordered state and are rich in raft-like components, they might correspond to rafts at least to some extent. Here we monitor the lateral order of biological membranes by characterizing DRMs from erythrocytes obtained with Brij-98, Brij-58, and TX-100 at 4?°C and 37?°C. All DRMs were enriched in cholesterol and contained the raft markers flotillin-2 and stomatin. However, sphingomyelin (SM) was only found to be enriched in TX-100-DRMs – a detergent that preferentially solubilizes the membrane inner leaflet – while Band 3 was present solely in Brij-DRMs. Electron paramagnetic resonance spectra showed that the acyl chain packing of Brij-DRMs was lower than TX-100-DRMs, providing evidence of their diverse lipid composition. Fatty acid analysis revealed that the SM fraction of the DRMs was enriched in lignoceric acid, which should specifically contribute to the resistance of SM to detergents. These results indicate that lipids from the outer leaflet, particularly SM, are essential for the formation of the liquid-ordered phase of DRMs. At last, the differential solubilization process induced by Brij-98 and TX-100 was monitored using giant unilamellar vesicles. This study suggests that Brij and TX-100-DRMs reflect different degrees of lateral order of the membrane microdomains. Additionally, Brij DRMs are composed by both inner and outer leaflet components, making them more physiologically relevant than TX-100-DRMs to the studies of membrane rafts.     

The topology of phosphatidylglycerol populations is essential for sustaining photosynthetic electron flow activities in thylakoid membranes

The transmembrane distribution of phosphatidylglycerol (PG) was determined in rightside-out (RO) and inside-out vesicles (IO) obtained by fragmentation of spinach thylakoids in a Yeda press, followed by partition in an aqueous dextran-polyethyleneglycol two-phase system. Using the phospholipase A(2) from porcine pancreas to digest selectively PG molecules in the outer monolayer (exposed to the incubation medium) of the membrane, we found the molar outside/inside distribution to be 70/30+/-5 in RO and 40/60+/-3 in IO. The transmembrane distribution of PG in IO was the opposite of that in intact thylakoids (molar ratio 58/42+/-3). The phospholipid population which sustained most of the uncoupled photosystem II electron flow activity was localized in the inner monolayer (exposed to the thylakoid lumen) of both thylakoid and RO membranes. In contrast, the activity in IO membranes was highly dependent on the PG population located in the outer monolayer. This finding brings the first direct demonstration of the dependence of the photosynthetic electron flow activity on the integrity of the inner topological pool of PG in the thylakoid membrane.     

Ultrastructure of cell wall and thylakoid membranes of the thermophilic cyanobacterium Synechococcus lividus under the influence of temperature shifts

The ultrastructure of the cell wall and the thylakoid membranes of the thermophilic cyanobacterium Synechococcus lividus was studied by freezefracture electron microscopy after temperature shifts. Different fracture faces of the outer, the cytoplasmic and the thylakoid membranes were demonstrated when the preparation-temperature was in the range of the optimal growth temperature at 52°C or after fixation at 52°C. In the outer membrane of the cell wall two fracture faces with holes and 7.5 nm intramembrane particles were detected. On both the outer (EF) and inner (PF) leaflet of the cytoplasmic membrane randomly distributed particles were demonstrated. The particle density on the PF-face was approx. three times that of the EF-face. The EF-face of the thylakoid membrane exposed rows of particles with an average diameter of 10 nm. The spacing between the particle rows was 35–50 nm. This regular particle arrangement on the EF-face was demonstrated only in a few cases. Mostly the intramembrane particles were distributed randomly on the thylakoid fracture faces. The particle density of thylakoids with a random distribution was approx. in the same range both on the EF-and PF-face. The EF-particles fall into four groups of 9,10,11, and 12.5 nm. The main particle class was the 10 nm class. The PF-face exposed smaller particles with two maxima at 8.5–9 nm and 10 nm. When Synechococcus lividus OH-53s was chilled to temperatures below 30–35°C before the freeze-etch preparation a phase transition took place after the temperature shift. On the fracture faces of the thylakoid and cytoplasmic membranes particle depleted areas occurred. The size of the areas were different in both membranes and dependent on the velocity of cooling. Contrary to Synechococcus lividus OH-53s in the mesophilic Synechococcus strain 6910 the phase transition point was 15°C. The lower phase transition point may be due to a higher content of unsaturated fatty acids.Dedicated to Prof. D. Peters (Hamburg) on the occasion of the 65th anniversary of his birthday     

Structural reorganization of thylakoid systems in response to heat treatment

The structural reorganization of pea thylakoid systems in response to osmotic shock in a wide range of temperatures (36–70°C) was studied. At temperatures 40–46°C, the configuration of thylakoid systems changed from a flattened to a nearly round, whereas thylakoids themselves remained compressed. The percentage of thylakoids stacked into grana at 44°C decreased from 71 % in the control to 40 % in experimental samples, reaching 59 % at 48°C. At 44°C and above, thylakoid systems ceased to respond to the osmotic shock by disordering, in contrast to what happened at lower temperatures (36–43°C) and in the control, and retained the configuration inherent in thylakoid systems at these temperatures. At 50°C and above, the packing of thylakoids in grana systems changed, and thylakoids formed extended strands of pseudograna. Simultaneously, single thylakoids formed a network of anastomoses through local fusions. At temperatures of 60–70°C, thylakoid systems appeared as spherical clusters of membrane vesicles with different degree of separation.This revised version was published online in March 2005 with corrections to the page numbers.     

Influence of thylakoid membrane lipids on the structure of aggregated light‐harvesting complexes of the diatom Thalassiosira pseudonana and the green alga Mantoniella squamata

The study investigated the effect of the thylakoid membrane lipids monogalactosyldiacylglycerol (MGDG), digalactosyldiacylglycerol (DGDG), sulphoquinovosyldiacylglycerol (SQDG) and phosphatidylglycerol (PG) on the structure of two algal light‐harvesting complexes (LHCs). In contrast to higher plants whose thylakoid membranes are characterized by an enrichment of the neutral galactolipids MGDG and DGDG, both the green alga Mantoniella squamata and the centric diatom Thalassiosira pseudonana contain membranes with a high content of the negatively charged lipids SQDG and PG. The algal thylakoids do not show the typical grana–stroma differentiation of higher plants but a regular arrangement. To analyze the effect of the membrane lipids, the fucoxanthin chlorophyll protein (FCP) complex of T. pseudonana and the LHC of M. squamata (MLHC) were prepared by successive cation precipitation using Triton X‐100 as detergent. With this method, it is possible to isolate LHCs with a reduced amount of associated lipids in an aggregated state. The results from 77 K fluorescence and photon correlation spectroscopy show that neither the neutral galactolipids nor the negatively charged lipids are able to significantly alter the aggregation state of the FCP or the MLHC. This is in contrast to higher plants where SQDG and PG lead to a strong disaggregation of the LHCII whereas MGDG and DGDG induce the formation of large macroaggregates. The results indicate that LHCs which are integrated into thylakoid membranes with a high amount of negatively charged lipids and a regular arrangement are less sensitive to lipid‐induced structural alterations than their counterparts in membranes enriched in neutral lipids with a grana–stroma differentiation.     

Glycoengineering of cyanobacterial thylakoid membranes for future studies on the role of glycolipids in photosynthesis

The lipid composition of thylakoid membranes is conserved from cyanobacteria to angiosperms. The predominating components are monogalactosyl- and digalactosyldiacylglycerol. In cyanobacteria, thylakoid membrane biosynthesis starts with the formation of monoglucosyldiacylglycerol which is C4-epimerized to the corresponding galactolipid, whereas in plastids monogalactosyldiacylglycerol is formed at the beginning. This suggests that galactolipids have specific functions in thylakoids. We wanted to investigate whether galactolipids can be replaced by glycosyldiacylglycerols with headgroups differing in their epimeric and anomeric details as well as the attachment point of the terminal hexose in diglycosyldiacylglycerols. For this purpose putative glycosyltransferase sequences were identified in databases to be used for functional expression in various host organisms. From 18 newly identified sequences, four turned out to encode glycosyltransferases catalyzing final steps in glycolipid biosynthesis: two alpha-glucosyltransferases, one beta-galactosyltransferase and one beta-glucosyltransferase. Their functional annotation was based on detailed structural characterization of the new glycolipids formed in the transformant hosts as well as on in vitro enzymatic assays. The expression of alpha-glucosyltransferases in the cyanobacterium Synechococcus resulted in the accumulation of the new alpha-galactosyldiacylglycerol which is ascribed to epimerization of the corresponding glucolipid. The expression of the beta-glucosyltransferase led to a high proportion of new beta-glucosyl-(1-->6)-beta-galactosyldiacylglycerol almost entirely replacing the native digalactosyldiacylglycerol. These results demonstrate that modifications of the glycolipid pattern in thylakoids are possible.     

Ultrastructure of the Thylakoid Membrane in Tomato Leaf Chloroplast Revealed by Liquid Helium Rapid-Freezing and Substitution-Fixation Method

The ultrastructure of the chloroplast in tomato leaves was studiedby a liquid helium rapid-freezing and substitution-fixationmethod. Distinct morphological differences were observed betweenthe ultrastructure of the thylakoid membrane fixed by this methodand the one fixed by the conventional chemical fixation method.The membrane was asymmetrical and consisted of triple-layeredleaflets. While the leaflet facing the stroma (outer leaflet)was very electron-dense, the one facing the thylakoid lumen(inner leaflet) was less electron-dense. The triple-layeredleaflets showed a structural difference between the grana thylakoidand the stroma thylakoid. In the grana region, where thylakoidmembranes are appressed, the part of outer leaflets were lookedlike fused or adhered to each other making one highly electron-denselayer composed of linings of particles (about 4 nm in diameter).The inner leaflet of the grana thylakoid membrane showed discontinuousstructure. This discontinuous structure was composed of a clusterof irregular particles (about 2–3 nm in diameter). Thesize of cluster varied from about 10nm to 16nm. The discontinuityof the inner leaflet, i.e. linings of the clusters at ratherconstant intervals, was not observed clearly in the stroma thylakoid.The liquid helium rapid-freezing and substitution-fixation methodcan be used to observe the details of protein complexes in thethylakoid membrane organization. (Received March 23, 1988; Accepted August 17, 1988)     

Polar lipid composition of chloroplast thylakoids isolated from leaves grown under different lighting conditions

The polar acyl lipid composition was determined for samples of chloroplast thylakoids isolated from Pisum sativum plants grown at light intensities of 50 and 300 E·m^-2·s^-1 and from Aesculus hippocastanum leaves taken from shade or sun environments. Lighting conditions had no major effect on lipid class composition except for a small increase in the amount of monogalactosyldiacylglycerol relative to other lipids in low compared with high light and shade compared with sun conditions. The thylakoids from low light and shade environments also had, relative to those from high light and sun conditions, a substantial decrease in the level of trans-hexadecenoic acid in phosphatidyglycerol. In parallel with this there were lower lipid to chlorophyll ratios, higher overall fatty acid unsaturation, lower chlorophyll a to b ratios and increased relative levels of light harvesting chlorophyll a/b polypeptides as expected for an increase in the degree of thylakoid appression. With this in mind, our results on lipid class composition and content of trans-hexadecenoic acid are discussed in the context of the lateral distribution of lipids within the plane of membrane.Abbreviations DGDG digalactosyldiacylglycerol - EDTA ethylenediaminetetraacetic acid - HEPES N-2-hydroxyethylpiperazine-N-2-ethanesulfonic acid - LHC light harvesting chlorophyll a/b - MGDG monogalactosyldiacylglycerol - MPL minor phospholipids - PS1 photosystem one - PS2 photosystem two - SDS sodium dodecyl sulphate - SL sulphoquinovosyldiacylglycerol     

Asymmetric distribution of phosphatidylethanolamine fatty acyl chains in the membrane of vesicular stomatitis virus

The membrane of vesicular stomatitis virus (VSV) contains two distinct pools of phosphatidylethanolamine molecules which reside in the inner and outer phospholipid monolayers, respectively. 36% of the total membrane phosphatidylethanolamine is found in the outer monolayer while 64% is found in the inner. The two pools of VSV phosphatidylethanolamine can be distinguished operationally by the fact that only outer phosphatidylethanolamine is reactive in intact virions with the membrane-impermeable reagent trinitrobenzenesulfonate (TNBS). We have made use of this property to separate inner from outer VSV phosphatidylethanolamine and to determine the fatty acyl chain compositions of the two phosphatidylethanolamine pools separately. The results show that compared to outer phosphatidylethanolamine, inner phosphatidylethanolamine molecules contain a significantly higher proportion of unsaturated fatty acyl chains. Furthermore, whereas the proportion of unsaturated fatty acyl chains was found to be quite similar at the 1 and 2 glycerol carbon atoms in inner phosphatidylethanolamine, a marked dissimilarity was observed in outer phosphatidylethanolamine; outer phosphatidylethanolamine was enriched in saturated fatty acyl chains at the 1 position and in unsaturated fatty acyl chains at the 2 position. The differential fatty acyl chain composition of inner compared to outer phosphatidylethanolamine indicates that rapid, random transmembrane migration (flip-flop) of phosphatidylethanolamine does not occur in the VSV membrane. The nature of the fatty acyl chain asymmetry observed in VSV phosphatidylethanolamine does not support the view that the     

31P-NMR observation of the temperature and glycerol induced non-lamellar phase formation in wheat thylakoid membranes

³¹P-NMR spectra at 162 MHz were used to monitor phase changes of wheat thylakoid membranes as a function of temperature. At room temperature the³¹P-NMR line was a superposition of anisotropic component characteristic of phospholipid lamellar phase and isotropic line due to inorganic phosphorus or small membrane vesicles arising as an effect of preparation. For temperatures higher than +35 °C an increase of the isotropic component occurs, which is irreversible as the sample is cooled. For the temperatures between +55 °C and +60 °C the presence of the hexagonal phase cylinders is suggested, as monitored by phosphorus lineshape. However, the addition of glycerol stimulates a formation of the isotropic phase. The effect of reconstitution of freeze-dried thylakoid membranes by addition of water or water-glycerol medium to the sample was examined. As lyophilizate was gradually diluted, the increase of isotropic line component was observed. For thylakoid membranes suspended in D₂O at the highest dilution examined, the line contribution due to small membrane fragments is not greater than 50%, but in presence of glycerol, this contribution could reach 70%. This suggests that the presence of glycerol increases the formation of the small membrane particles as the thylakoid membrane is reconstituted from lyophilizate. The wheat thylakoid membranes reconstituted from lyophilizate show, in comparison to native membranes, the increased contribution of small membrane vesicles. Moreover, the³¹P -NMR spectra suggest the appearance of the hexagonal phase cylinders even at +50 °C.Abbreviations DGDG digalactosyldiacylglycerol - DLPC dilinoleoyl phosphatidylcholine - DLPE dilinoleoyl phosphatidylethanolamine - EDTA ethylenediamine-tetraacetic acid - MGDG monogalactosyldiacylglycerol - NMR nuclear magnetic resonance - PC phosphatidylcholine - PG phosphatidylglycerol - PSII photosystem II - TGDG trigalactosyldiacylglycerol - Tris Tris-(hydroxymethyl)-aminomethan - S/N signal to noise ratio