Nejvíce citovaný článek - PubMed ID 35351472
Structural and functional roles of non-bilayer lipid phases of chloroplast thylakoid membranes and mitochondrial inner membranes
Thylakoid membranes (TMs) of oxygenic photosynthetic organisms are flat membrane vesicles, which form highly organised, interconnected membrane networks. In vascular plants, they are differentiated into stacked and unstacked regions, the grana and stroma lamellae, respectively; they are densely packed with protein complexes performing the light reactions of photosynthesis and generating a proton motive force (pmf). The maintenance of pmf and its utilisation for ATP synthesis requires sealing the TMs at their highly curved regions (CRs). These regions are devoid of chlorophyll-containing proteins but contain the curvature-inducing CURVATURE THYLAKOID1 (CURT1) proteins and are enriched in lipids. Because of the highly curved nature of this region, at the margins of grana and stroma TMs, the molecular organisation of lipid molecules is likely to possess distinct features compared to those in the major TM domains. To clarify this question, we isolated CR fractions from Spinacia oleracea and, using BN-PAGE and western blot analysis, verified that they are enriched in CURT1 proteins and in lipids. The lipid phase behaviour of these fractions was fingerprinted with 31P-NMR spectroscopy, which revealed that the bulk lipid molecules assume a non-bilayer, isotropic lipid phase. This finding underpins the importance of the main, non-bilayer lipid species, monogalactosyldiacylglycerol, of TMs in their self-assembly and functional activity.
- Klíčová slova
- 31P‐NMR, CURT1 protein, granum margin, non‐bilayer lipid phase, thylakoid membrane,
- MeSH
- lipidy * chemie MeSH
- rostlinné proteiny metabolismus MeSH
- Spinacia oleracea * metabolismus MeSH
- tylakoidy * metabolismus chemie MeSH
- Publikační typ
- časopisecké články MeSH
- Názvy látek
- lipidy * MeSH
- rostlinné proteiny MeSH
In oxygenic photosynthetic organisms, the light reactions are performed by protein complexes embedded in the lipid bilayer of thylakoid membranes (TMs). The organization of the bulk lipid molecules into bilayer structures provide optimal conditions for the build-up of the proton motive force (pmf) and its utilization for ATP synthesis. However, the lipid composition of TMs is dominated by the non-bilayer lipid species monogalactosyl diacylglycerol (MGDG), and functional plant TMs, besides the bilayer, contain large amounts of non-bilayer lipid phases. Bulk lipids have been shown to be associated with lumenal, stromal-side and marginal-region proteins and proposed to play roles in the self-assembly and photoprotection of the photosynthetic machinery. Furthermore, it has recently been pointed out that the generation and utilization of pmf for ATP synthesis according to the 'protet' or protonic charge transfer model Kell (Biochim Biophys Acta Bioenerg 1865(4):149504, 2024), requires high MGDG content Garab (Physiol Plant 177(2):e70230, 2025). In this study, to gain better insight into the structural and functional roles of MGDG, we employed all atom and coarse-grained molecular dynamics simulations to explore how temperature, hydration levels and varying MGDG concentrations affect the structural and dynamic properties of bilayer membranes constituted of plant thylakoid lipids. Our findings reveal that MGDG promotes increased membrane fluidity and dynamic fluctuations in membrane thickness. MGDG-rich stacked bilayers spontaneously formed inverted hexagonal phases; these transitions were enhanced at low hydration levels and at elevated but physiologically relevant temperatures. It can thus be inferred that MGDG plays important roles in heat and drought stress mechanisms.
- Klíčová slova
- Dehydration, Inverted hexagonal phase, MGDG, Non-bilayer lipid, Thylakoid membranes,
- MeSH
- fotosyntéza MeSH
- galaktolipidy metabolismus chemie MeSH
- lipidové dvojvrstvy metabolismus chemie MeSH
- simulace molekulární dynamiky * MeSH
- teplota MeSH
- tylakoidy * metabolismus chemie MeSH
- Publikační typ
- časopisecké články MeSH
- Názvy látek
- galaktolipidy MeSH
- lipidové dvojvrstvy MeSH
- monogalactosyldiacylglycerol MeSH Prohlížeč
The light reactions of oxygenic photosynthesis are performed by protein complexes embedded in the lipid bilayer of thylakoid membranes (TMs). Bilayers provide optimal conditions for the build-up of the proton motive force (pmf) and ATP synthesis. However, functional plant TMs, besides the bilayer, contain an inverted hexagonal (HII) phase and isotropic phases, a lipid polymorphism due to their major, non-bilayer lipid species, monogalactosyldiacylglycerol (MGDG). The lipid phase behavior of TMs is explained within the framework of the Dynamic Exchange Model (DEM), an extension of the fluid-mosaic model. DEM portrays the bilayer phase as inclusions between photosynthetic supercomplexes - characterized by compromised membrane impermeability and restricted sizes inflicted by the segregation propensity of lipid molecules, safe-guarding the high protein density of TMs. Isotropic phases mediate membrane fusions and are associated with the lumenal lipocalin-like enzyme, violaxanthin de-epoxidase. Stromal-side proteins surrounded by lipids give rise to the HII phase. These features instigate experimentally testable hypotheses: (i) non-bilayer phases mediate functional sub-compartmentalization of plant chloroplasts - a quasi-autonomous energization and ATP synthesis of each granum-stroma TM assembly; and (ii) the generation and utilization of pmf depend on hydrated protein networks and proton-conducting pathways along membrane surfaces - rather than on strict impermeability of the bilayer.
It has been thoroughly documented, by using 31P-NMR spectroscopy, that plant thylakoid membranes (TMs), in addition to the bilayer (or lamellar, L) phase, contain at least two isotropic (I) lipid phases and an inverted hexagonal (HII) phase. However, our knowledge concerning the structural and functional roles of the non-bilayer phases is still rudimentary. The objective of the present study is to elucidate the origin of I phases which have been hypothesized to arise, in part, from the fusion of TMs (Garab et al. 2022 Progr Lipid Res 101,163). We take advantage of the selectivity of wheat germ lipase (WGL) in eliminating the I phases of TMs (Dlouhý et al. 2022 Cells 11: 2681), and the tendency of the so-called BBY particles, stacked photosystem II (PSII) enriched membrane pairs of 300-500 nm in diameter, to form large laterally fused sheets (Dunahay et al. 1984 BBA 764: 179). Our 31P-NMR spectroscopy data show that BBY membranes contain L and I phases. Similar to TMs, WGL selectively eliminated the I phases, which at the same time exerted no effect on the molecular organization and functional activity of PSII membranes. As revealed by sucrose-density centrifugation, magnetic linear dichroism spectroscopy and scanning electron microscopy, WGL disassembled the large laterally fused sheets. These data provide direct experimental evidence on the involvement of I phase(s) in the fusion of stacked PSII membrane pairs, and strongly suggest the role of non-bilayer lipids in the self-assembly of the TM system.
- Klíčová slova
- 31P-NMR spectroscopy; BBY membrane, Linear dichroism spectroscopy, Membrane fusion; non-bilayer lipids, Wheat germ lipase,
- MeSH
- fotosystém II (proteinový komplex) * metabolismus MeSH
- fúze membrán fyziologie MeSH
- lipidy chemie MeSH
- magnetická rezonanční spektroskopie MeSH
- tylakoidy * metabolismus MeSH
- Publikační typ
- časopisecké články MeSH
- Názvy látek
- fotosystém II (proteinový komplex) * MeSH
- lipidy MeSH
In our earlier works, we have shown that the rate-limiting steps, associated with the dark-to-light transition of Photosystem II (PSII), reflecting the photochemical activity and structural dynamics of the reaction center complex, depend largely on the lipidic environment of the protein matrix. Using chlorophyll-a fluorescence transients (ChlF) elicited by single-turnover saturating flashes, it was shown that the half-waiting time (Δτ 1/2) between consecutive excitations, at which 50% of the fluorescence increment was reached, was considerably larger in isolated PSII complexes of Thermostichus (T.) vulcanus than in the native thylakoid membrane (TM). Further, it was shown that the addition of a TM lipid extract shortened Δτ 1/2 of isolated PSII, indicating that at least a fraction of the 'missing' lipid molecules, replaced by detergent molecules, caused the elongation of Δτ 1/2. Here, we performed systematic experiments to obtain information on the nature of TM lipids that are capable of decreasing Δτ 1/2. Our data show that while all lipid species shorten Δτ 1/2, the negatively charged lipid phosphatidylglycerol appears to be the most efficient species - suggesting its prominent role in determining the structural dynamics of PSII reaction center.
- Klíčová slova
- chlorophyll-a fluorescence, core complex of photosystem II, rate-limiting step, structural dynamics, thylakoid lipids, waiting time,
- Publikační typ
- časopisecké články MeSH
To express my appreciation of Győző Garab, I describe below our association over the years, followed by a brief description of his fundamental research in the area of biophysics of photosynthesis.
- Klíčová slova
- Hartmut Lichtenthaler, Hartmut Michel, Jan Anderson, Paul Mathis, Rajni Govindjee, Reto Strasser, Wolfgang Junge,
- Publikační typ
- časopisecké články MeSH
Thylakoid membranes are energy-converting membranes with a unique lipid composition. Though the membranes are primarily composed of proteins, their photosynthetic function is strongly influenced by the lipid constituents. Here we characterize, with molecular dynamics (MD) simulations, lipid bilayers with compositions representative of plant thylakoid membranes. We determine, in a wide range of temperatures, the physical parameters of the model membranes which are relevant for the photosynthetic function. We found a marked impact of temperature on membrane permeability due to a combination of increased compressibility and curvature of the membrane at elevated temperatures. With increasing temperatures, we observed increasingly smeared transmembrane density profiles of the membrane forming lipid headgroups predicting increased membrane flexibility. The diffusion coefficient of the lipids increased with temperature without apparent specificity for lipid species. Instead of a comprehensive experimental dataset in the relevant temperature range, we quantitatively compared and validated our MD results with MD simulations on a dipalmitoylphosphatidylcholine model system.
- Klíčová slova
- mechanical properties, molecular dynamics, permeability, thylakoid,
- Publikační typ
- časopisecké články MeSH
It is well established that plant thylakoid membranes (TMs), in addition to a bilayer, contain two isotropic lipid phases and an inverted hexagonal (HII) phase. To elucidate the origin of non-bilayer lipid phases, we recorded the 31P-NMR spectra of isolated spinach plastoglobuli and TMs and tested their susceptibilities to lipases and proteases; the structural and functional characteristics of TMs were monitored using biophysical techniques and CN-PAGE. Phospholipase-A1 gradually destroyed all 31P-NMR-detectable lipid phases of isolated TMs, but the weak signal of isolated plastoglobuli was not affected. Parallel with the destabilization of their lamellar phase, TMs lost their impermeability; other effects, mainly on Photosystem-II, lagged behind the destruction of the original phases. Wheat-germ lipase selectively eliminated the isotropic phases but exerted little or no effect on the structural and functional parameters of TMs-indicating that the isotropic phases are located outside the protein-rich regions and might be involved in membrane fusion. Trypsin and Proteinase K selectively suppressed the HII phase-suggesting that a large fraction of TM lipids encapsulate stroma-side proteins or polypeptides. We conclude that-in line with the Dynamic Exchange Model-the non-bilayer lipid phases of TMs are found in subdomains separated from but interconnected with the bilayer accommodating the main components of the photosynthetic machinery.
- Klíčová slova
- 31P-NMR spectroscopy, lipid polymorphism, lipocalins, membrane fusion, membrane models, non-bilayer lipids, plastoglobuli, structural and functional plasticity, thylakoid membrane,
- MeSH
- lipasa metabolismus MeSH
- lipidové dvojvrstvy * metabolismus MeSH
- magnetická rezonanční spektroskopie MeSH
- proteasy metabolismus MeSH
- tylakoidy * metabolismus MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
- Názvy látek
- lipasa MeSH
- lipidové dvojvrstvy * MeSH
- proteasy MeSH
Alcohols are a part of cellular metabolism, but their physiological roles are not well understood. We investigated the effects of short-chain alcohols on Daphnia pulex and model membranes mimicking the lipid composition of eukaryotic inner mitochondrial membranes. We also studied the synergistic effects of alcohols with the bee venom membrane-active peptide, melittin, which is structurally similar to endogenous membrane-active peptides. The alcohols, from ethanol to octanol, gradually decreased the heart rate and the mitochondrial ATP synthesis of daphnia; in contrast, in combination with melittin, which exerted no sizeable effect, they gradually increased both the heart rate and the ATP synthesis. Lipid packing and the order parameter of oriented films, monitored by EPR spectroscopy of the spin-labeled probe 5-doxylstrearic acid, revealed gradual alcohol-assisted bilayer to non-bilayer transitions in the presence of melittin; further, while the alcohols decreased, in combination with melittin they increased the order parameter of the film, which is attributed to the alcohol-facilitated association of melittin with the membrane. A 1H-NMR spectroscopy of the liposomes confirmed the enhanced induction of a non-bilayer lipid phase that formed around the melittin, without the permeabilization of the liposomal membrane. Our data suggest that short-chain alcohols, in combination with endogenous peptides, regulate protein functions via modulating the lipid polymorphism of membranes.
- Klíčová slova
- 1H-NMR, ERP of spin probes, alcohols, heart rate, melittin, mitochondrial ATP production, non-bilayer structures,
- MeSH
- adenosintrifosfát MeSH
- alkoholy farmakologie MeSH
- lipidy MeSH
- liposomy MeSH
- melitten * chemie metabolismus farmakologie MeSH
- včelí jedy * farmakologie MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
- Názvy látek
- adenosintrifosfát MeSH
- alkoholy MeSH
- lipidy MeSH
- liposomy MeSH
- melitten * MeSH
- včelí jedy * MeSH
