Thermal stability of trimeric light-harvesting chlorophyll a/b complex (LHCIIb) in liposomes of thylakoid lipids

The major light-harvesting chlorophyll a/b complex (LHCIIb) of photosystem (PS) II functions by harvesting light energy and by limiting and balancing the energy flow directed towards the PSI and PSII reaction centers. The complex is predominantly trimeric; however, the monomeric form may play a role in one or several of the regulatory functions of LHCIIb. In this work the dissociation temperature was measured of trimeric LHCIIb isolated from Pisum thylakoids and inserted into liposomes made of various combinations of thylakoid lipids at various protein densities. Dissociation was measured by monitoring a trimer-specific circular dichroism signal in the visible range. The LHCIIb density in the membrane significantly affected the trimer dissociation temperature ranging from 70 °C at an LHCIIb concentration comparable to or higher than the one in thylakoid grana, to 65 °C at the density estimated in stromal lamellae. Omitting one thylakoid lipid from the liposomes had virtually no effect on the thermal trimer stability in most cases except when digalactosyl diacylglycerol (DGDG) was omitted which caused a drop in the apparent dissociation temperature by 2 °C. In liposomes containing only one lipid species, DGDG and, even more so, monogalactosyl diacylglycerol (MGDG) increased the thermal stability of LHCIIb trimers whereas phosphatidyl diacylglycerol (PG) significantly decreased it. The lateral pressure exerted by the non-bilayer lipid MGDG did not significantly influence LHCII trimer stability.     

Cold acclimation of the Arabidopsis dgd1 mutant results in recovery from photosystem I-limited photosynthesis

We compared the thylakoid membrane composition and photosynthetic properties of non- and cold-acclimated leaves from the dgd1 mutant (lacking >90% of digalactosyl-diacylglycerol; DGDG) and wild type (WT) Arabidopsis thaliana. In contrast to warm grown plants, cold-acclimated dgd1 leaves recovered pigment-protein pools and photosynthetic function equivalent to WT. Surprisingly, this recovery was not correlated with an increase in DGDG. When returned to warm temperatures the severe dgd1 mutant phenotype reappeared. We conclude that the relative recovery of photosynthetic activity at 5 degrees C resulted from a temperature/lipid interaction enabling the stable assembly of PSI complexes in the thylakoid.     

Lack of digalactosyldiacylglycerol increases the sensitivity of Synechocystis sp. PCC 6803 to high light stress

The physiological role of digalactosyldiacylglycerol (DGDG) in photosynthesis was examined using a dgdA mutant of Synechocystis sp. PCC 6803 that is defective in the biosynthesis of DGDG. The dgdA mutant cells showed normal growth under low light (LL) conditions. However, their growth was retarded under high light (HL) conditions and under Ca²⁺- and/or Cl⁻-limited conditions compared to wild-type cells. The retardation in growth of the mutant cells was recovered by exogenous supply of DGDG in the growth medium. The dgdA mutant showed increased sensitivity to photoinhibition. Although both photodamage and repair processes of photosynthesis were affected, the repair process was more severely affected than the photodamage process, suggesting that DGDG plays an important role in the photosynthetic repair cycle.     

Differential changes in galactolipid and phospholipid species in soybean leaves and roots under nitrogen deficiency and after nodulation

The availability of nitrogen (N) to plants has a profound impact on carbohydrate and protein metabolism, but little is known about its effect on membrane lipid species. This study examines the changes in galactolipid and phospholipid species in soybean as affected by the availability of N, either supplied to soil or obtained through Bradyrhizobium japonicum nodulation. When N was limited in soil, the content of galactolipids, monogalactosyldiacylglycerol (MGDG) and digalactosyldiacyglycerol (DGDG), decreased drastically in leaves, while a smaller decrease of DGDG was observed in roots. In both leaves and roots, the overall content of different phospholipid classes was largely unchanged by N limitation, although some individual phospholipid molecular species did display significant changes. Nodulation with Bradyrhizobium of soybean grown in N-deficient soil resulted in a large increase in levels of plastidic lipid classes, MGDG, DGDG, and phosphatidylglycerol, along with smaller increases in non-plastidic phospholipids in leaves. Nodulation also led to higher levels of phospholipids in roots without changes in root levels of MGDG and DGDG. Overall, N availability alters lipid content more in leaves than roots and more in galactolipids than phospholipids. Increased N availability leads to increased galactolipid accumulation in leaves, regardless of whether N is supplied from the soil or symbiotic fixation.     

Red (hot) algae: modulation of mono- and digalactosyldiacylglycerol-associated fatty acids of Polysiphonia sp. and Porphyridium sp. in response to growth temperature

ABSTRACT

There is a dearth of surveys examining the direct effects of temperature on red algal galactolipids, and none which examine regiochemistry modulation with respect to growth temperature. Therefore, forms of monogalactosyldiacylglycerol (MGDG) and digalactosyldiacylglycerol (DGDG), the two most commonly found galactolipids in chloroplast membranes, were determined in two model red algae, Polysiphonia sp. and Porphyridium sp., via positive-ion electrospray ionization/mass spectrometry (ESI/MS) and ESI/MS/MS. We sought to compare modulation of galactolipid forms in response to growth temperature between these two red algae and selected descendants with red algal plastid ancestry, and have proposed the following hypothesis: Polysiphonia sp. and Porphyridium sp. would modulate desaturations in the sn-2 position in accordance with previously examined descendant organisms. It was observed that both red algae produced C₂₀/C₁₆ (sn-1/sn-2 regiochemistry) and C₂₀/C₂₀ forms of MGDG and DGDG as their most abundant galactolipids under two growth temperatures, 20°C and 30°C. Furthermore, temperature-induced modulation of the major forms of MGDG and DGDG was more complex than what has been observed previously in selected representatives of red algal plastid ancestry. Porphyridium sp. modulated levels of desaturation in the sn-1 position of C₂₀/C₁₆ forms of MGDG and DGDG and in the sn-1 and sn-2 positions of C₂₀/C₂₀ forms of MGDG and DGDG. Polysiphonia sp. displayed trends suggesting it modulates levels of desaturation in the sn-1 and sn-2 positions of C₂₀/C₂₀ forms of MGDG and DGDG, thus indicating a different approach to regulating plastid membrane fluidity from that which has been observed in algae with secondary, red algae-derived plastids.     

Conservation of lipid functions in cytochrome bc complexes

Lipid binding sites and properties are compared in two sub-families of hetero-oligomeric membrane protein complexes known to have similar functions in order to gain further understanding of the role of lipid in the function, dynamics, and assembly of these complexes. Using the crystal structure information for both complexes, we compared the lipid binding properties of the cytochrome b₆f and bc₁ complexes that function in photosynthetic and respiratory membrane energy transduction. Comparison of lipid and detergent binding sites in the b₆f complex with those in bc₁ shows significant conservation of lipid positions. Seven lipid binding sites in the cyanobacterial b₆f complex overlap three natural sites in the Chlamydomonas reinhardtii algal complex and four sites in the yeast mitochondrial bc₁ complex. The specific identity of lipids is different in b₆f and bc₁ complexes: b₆f contains sulfoquinovosyldiacylglycerol, phosphatidylglycerol, phosphatidylcholine, monogalactosyldiacylglycerol, and digalactosyldiacylglycerol, whereas cardiolipin, phosphatidylethanolamine, and phosphatidic acid are present in the yeast bc₁ complex. The lipidic chlorophyll a and β-carotene (β-car) in cyanobacterial b₆f, as well as eicosane in C. reinhardtii, are unique to the b₆f complex. Inferences of lipid binding sites and functions were supported by sequence, interatomic distance, and B-factor information on interacting lipid groups and coordinating amino acid residues. The lipid functions inferred in the b₆f complex are as follows: (i) substitution of a transmembrane helix by a lipid and chlorin ring, (ii) lipid and β-car connection of peripheral and core domains, (iii) stabilization of the iron-sulfur protein transmembrane helix, (iv) n-side charge and polarity compensation, and (v) β-car-mediated super-complex with the photosystem I complex.     

Phylogeny of galactolipid synthase homologs together with their enzymatic analyses revealed a possible origin and divergence time for photosynthetic membrane biogenesis

The photosynthetic membranes of cyanobacteria and chloroplasts of higher plants have remarkably similar lipid compositions. In particular, thylakoid membranes of both cyanobacteria and chloroplasts are composed of galactolipids, of which monogalactosyldiacylglycerol (MGDG) is the most abundant, although MGDG biosynthetic pathways are different in these organisms. Comprehensive phylogenetic analysis revealed that MGDG synthase (MGD) homologs of filamentous anoxygenic phototrophs Chloroflexi have a close relationship with MGDs of Viridiplantae (green algae and land plants). Furthermore, analyses for the sugar specificity and anomeric configuration of the sugar head groups revealed that one of the MGD homologs exhibited a true MGDG synthetic activity. We therefore presumed that higher plant MGDs are derived from this ancestral type of MGD genes, and genes involved in membrane biogenesis and photosystems have been already functionally associated at least at the time of Chloroflexi divergence. As MGD gene duplication is an important event during plastid evolution, we also estimated the divergence time of type A and B MGDs. Our analysis indicated that these genes diverged -323 million years ago, when Spermatophyta (seed plants) were appearing. Galactolipid synthesis is required to produce photosynthetic membranes; based on MGD gene sequences and activities, we have proposed a novel evolutionary model that has increased our understanding of photosynthesis evolution.     

The influence of phase transitions in phosphatidylethanolamine models on the activity of violaxanthin de-epoxidase

In the present study, the influence of the phospholipid phase state on the activity of the xanthophyll cycle enzyme violaxanthin de-epoxidase (VDE) was analyzed using different phosphatidylethanolamine species as model lipids. By using ³¹P NMR spectroscopy, differential scanning calorimetry and temperature dependent enzyme assays, VDE activity could directly be related to the lipid structures the protein is associated with. Our results show that the gel (L_β) to liquid-crystalline (L_α) phase transition in these single lipid component systems strongly enhances both the solubilization of the xanthophyll cycle pigment violaxanthin in the membrane and the activity of the VDE. This phase transition has a significantly stronger impact on VDE activity than the transition from the L_α to the inverted hexagonal (H_II) phase. Especially at higher temperatures we found increased VDE reaction rates in the presence of the L_α phase compared to those in the presence of H_II phase forming lipids. Our data furthermore imply that the H_II phase is better suited to maintain high VDE activities at lower temperatures.     

The development of a new adjuvant lipid-saponin complex and its use at experimental immunization by bacterial antigen

Immunostimulating complexes (ISCOM) have been modified by replacement of structural components of ISCOM; phosphatidylcholine has been replaced, by glycolipid of monogalactosyldiacylglycerol from sea macrophytes, saponin QuilA has been replaced by triterpene glycoside, Cucumarioside A₂-2. The resultant complex includes morphological structures of two types: the ISCOM-like structures with the characteristic morphology and sizes and also the tubular structures with diameter of about 40 nm and length of 150–400 nm. We have defined these structures as TI-complexes. These TI-complexes exhibit considerably lower toxicity than ISCOM. They may include an amphiphilic protein antigen and provide immunoadjuvant effect during experimental vaccination. Under conditions of the experimental immunization of mice with a weak immunogen (subunit membrane pore protein from Y. pseudotuberculosis), TI-complexes with antigen provided stronger humoral immune response to antigen than the complexes of porin with classical ISCOM, liposomes and Freund’s adjuvant. Thus, the TI-complexes represent a new type of adjuvant carriers for antigens.     

Glyco- and sphingophosphonolipids from the medusa Phyllorhiza punctata: NMR and ESI-MS/MS fingerprints

The medusa Phyllorhiza punctata has been found in Brazilian waters where it is an exotic species, having arrived in ballasts from the Indo-Pacific Ocean in the general region of North Australia and Indonesia. Fatty acids of the intact animal and its component umbrella, oral arms, and mucus were identified. Two different groups of glycolipids and a sphingolipid were isolated by silica-gel column chromatography and characterized using GC-MS, ESI-MS, 1D, 2D (13)C, (1)H and (31)P NMR spectroscopy. They were sulfoquinovosyldiacylglycerol (SQDG), monogalactosyldiacylglycerol (MGDG), and ceramide aminoethylphosphonate (CAEP). The CAEP long chain base (LCB) and its polar head group (PHG) formed by partial hydrolysis, were analyzed by ESI-MS/MS. The probable origin of MGDG and SQDG in the jellyfish is the result of an endosymbiotic association with a microalga of the Dinoflagellate group, since these lipids are commonly found in photosynthetic membranes.