Common Phospholipids Research Articles

Nonspecific phospholipases C3 and C4 interact with PIN-FORMED2 to regulate growth and tropic responses in Arabidopsis.

The dynamic changes in membrane phospholipids affect membrane biophysical properties and cell signaling, thereby influencing numerous biological processes. Nonspecific phospholipase C (NPC) enzymes hydrolyze common phospholipids to release diacylglycerol (DAG), which is converted to phosphatidic acid (PA) and other lipids. In this study, 2 Arabidopsis (Arabidopsis thaliana) tandemly arrayed genes, NPC3 and NPC4, were identified as critical factors modulating auxin-controlled plant growth and tropic responses. Moreover, NPC3 and NPC4 were shown to interact with the auxin efflux transporter PIN-FORMED2 (PIN2). The loss of NPC3 and NPC4 enhanced the endocytosis and vacuolar degradation of PIN2, which disrupted auxin gradients and slowed gravitropic and halotropic responses. Furthermore, auxin-triggered activation of NPC3 and NPC4 is required for the asymmetric PA distribution that controls PIN2 trafficking dynamics and auxin-dependent tropic responses. Collectively, our study reveals an NPC-derived PA signaling pathway in Arabidopsis auxin fluxes that is essential for fine-tuning the balance between root growth and environmental responses.

Comprehensive analysis of the composition of the major phospholipids during the asexual life cycle of the filamentous fungus Aspergillus nidulans

Filamentous fungi undergo significant cellular morphological changes during their life cycle. It has recently been reported that deletions of genes that are involved in phospholipid synthesis led to abnormal hyphal morphology and differentiation in filamentous fungi. Although these results suggest the importance of phospholipid balance in their life cycle, comprehensive analyses of cellular phospholipids are limited. Here, we performed lipidomic analysis of A. nidulans during morphological changes in a liquid medium and of colonies on a solid medium. We observed that the phospholipid composition and transcription of the genes involved in phospholipid synthesis changed dynamically during the life cycle. Specifically, the levels of phosphatidylethanolamine, and highly unsaturated phospholipids increased during the establishment of polarity. Furthermore, we demonstrated that the phospholipid composition in the hyphae at colony margins is similar to that during conidial germination. Furthermore, we demonstrated that common and characteristic phospholipid changes occurred during germination in A. nidulans and A. oryzae, and that species-specific changes also occurred. These results suggest that the exquisite regulation of phospholipid composition is crucial for the growth and differentiation of filamentous fungi.

Bacterial catabolism of membrane phospholipids links marine biogeochemical cycles.

In marine systems, the availability of inorganic phosphate can limit primary production leading to bacterial and phytoplankton utilization of the plethora of organic forms available. Among these are phospholipids that form the lipid bilayer of all cells as well as released extracellular vesicles. However, information on phospholipid degradation is almost nonexistent despite their relevance for biogeochemical cycling. Here, we identify complete catabolic pathways for the degradation of the common phospholipid headgroups phosphocholine (PC) and phosphorylethanolamine (PE) in marine bacteria. Using Phaeobacter sp. MED193 as a model, we provide genetic and biochemical evidence that extracellular hydrolysis of phospholipids liberates the nitrogen-containing substrates ethanolamine and choline. Transporters for ethanolamine (EtoX) and choline (BetT) are ubiquitous and highly expressed in the global ocean throughout the water column, highlighting the importance of phospholipid and especially PE catabolism in situ. Thus, catabolic activation of the ethanolamine and choline degradation pathways, subsequent to phospholipid metabolism, specifically links, and hence unites, the phosphorus, nitrogen, and carbon cycles.

The LPA-CDK5-tau pathway mediates neuronal injury in an in vitro model of ischemia-reperfusion insult

Lysophosphatidic acid (LPA) is a common glycerol phospholipid and an important extracellular signaling molecule. LPA binds to its receptors and mediates a variety of biological effects, including the pathophysiological process underlying ischemic brain damage and traumatic brain injury. However, the molecular mechanisms mediating the pathological role of LPA are not clear. Here, we found that LPA activates cyclin-dependent kinase 5 (CDK5). CDK5 phosphorylates tau, which leads to neuronal cell death. Inhibition of LPA production or blocking its receptors reduced the abnormal activation of CDK5 and phosphorylation of tau, thus reversing the death of neurons. Our data indicate that the LPA-CDK5-Tau pathway plays an important role in the pathophysiological process after ischemic stroke. Inhibiting the LPA pathway may be a potential therapeutic target for treating ischemic brain injury.

Characterization of the Glehnia littoralis Non-specific Phospholipase C Gene GlNPC3 and Its Involvement in the Salt Stress Response.

Glehnia littoralis is a medicinal halophyte that inhabits sandy beaches and has high ecological and commercial value. However, the molecular mechanism of salt adaptation in G. littoralis remains largely unknown. Here, we cloned and identified a non-specific phospholipase C gene (GlNPC3) from G. littoralis, which conferred lipid-mediated signaling during the salt stress response. The expression of GlNPC3 was induced continuously by salt treatment. Overexpression of GlNPC3 in Arabidopsis thaliana increased salt tolerance compared to wild-type (WT) plants. GlNPC3-overexpressing plants had longer roots and higher fresh and dry masses under the salt treatment. The GlNPC3 expression pattern revealed that the gene was expressed in most G. littoralis tissues, particularly in roots. The subcellular localization of GlNPC3 was mainly at the plasma membrane, and partially at the tonoplast. GlNPC3 hydrolyzed common membrane phospholipids, such as phosphotidylserine (PS), phosphoethanolamine (PE), and phosphocholine (PC). In vitro enzymatic assay showed salt-induced total non-specific phospholipase C (NPC) activation in A. thaliana GlNPC3-overexpressing plants. Plant lipid profiling showed a significant change in the membrane-lipid composition of A. thaliana GlNPC3-overexpressing plants compared to WT after the salt treatment. Furthermore, downregulation of GlNPC3 expression by virus-induced gene silencing in G. littoralis reduced the expression levels of some stress-related genes, such as SnRK2, P5SC5, TPC1, and SOS1. Together, these results indicated that GlNPC3 and GlNPC3-mediated membrane lipid change played a positive role in the response of G. littoralis to a saline environment.

TTAPE-Me dye is not selective to cardiolipin and binds to common anionic phospholipids nonspecifically

Identification, visualization, and quantitation of cardiolipin (CL) in biological membranes is of great interest because of the important structural and physiological roles of this lipid. Selective fluorescent detection of CL using noncovalently bound fluorophore 1,1,2,2-tetrakis[4-(2-trimethylammonioethoxy)-phenylethene (TTAPE-Me) has been recently proposed. However, this dye was only tested on wild-type mitochondria or liposomes containing negligible amounts of other anionic lipids, such as phosphatidylglycerol (PG) and phosphatidylserine (PS). No clear preference of TTAPE-Me for binding to CL compared to PG and PS was found in our experiments on artificial liposomes, Escherichia coli inside-out vesicles, or Saccharomyces cerevisiae mitochondria in vitro or in situ, respectively. The shapes of the emission spectra for these anionic phospholipids were also found to be indistinguishable. Thus, TTAPE-Me is not suitable for detection, visualization, and localization of CL in the presence of other anionic lipids present in substantial physiological amounts. Our experiments and complementary molecular dynamics simulations suggest that fluorescence intensity of TTAPE-Me is regulated by dynamic equilibrium between emitting dye aggregates, stabilized by unspecific but thermodynamically favorable electrostatic interactions with anionic lipids, and nonemitting dye monomers. These results should be taken into consideration when interpreting past and future results of CL detection and localization studies with this probe in vitro and in vivo. Provided methodology emphasizes minimal experimental requirements, which should be considered as a guideline during the development of novel lipid-specific probes.

Spontaneous Formation of Stable Vesicles and Vesicle Gels in Polar Organic Solvents.

The self-assembly of lipids into nanoscale vesicles (liposomes) is routinely accomplished in water. However, reports of similar vesicles in polar organic solvents like glycerol, formamide, and ethylene glycol (EG) are scarce. Here, we demonstrate the formation of nanoscale vesicles in glycerol, formamide, and EG using the common phospholipid lecithin (derived from soy). The samples we study are simple binary mixtures of lecithin and the solvent, with no additional cosurfactants or salt. Lecithin dissolves readily in the solvents and spontaneously gives rise to viscous fluids at low lipid concentrations (∼2-4%), with structures ∼200 nm detected by dynamic light scattering. At higher concentrations (>10%), lecithin forms clear gels that are strongly birefringent at rest. Dynamic rheology confirms the elastic response of gels, with their elastic modulus being ∼20 Pa at ∼10% lipid. Images from cryo-scanning electron microscopy (cryo-SEM) indicate that concentrated samples are "vesicle gels," where multilamellar vesicles (MLVs, also called "onions"), with diameters between 50 and 600 nm, are close-packed across the sample volume. This structure can explain both the elastic rheology as well as the static birefringence of the samples. The discovery of vesicles and vesicle gels in polar solvents widens the scope of systems that can be created by self-assembly. Interestingly, it is much easier to form vesicles in polar solvents than in water, and the former are stable indefinitely, whereas the latter tend to aggregate or coalesce over time. The stability is attributed to refractive index-matching between lipid bilayers and the solvents, i.e., these vesicles are relatively "invisible" and thus experience only weak attractions. The ability to use lipids (which are "green" or eco-friendly molecules derived from renewable natural sources) to thicken and form gels in polar solvents could also prove useful in a variety of areas, including cosmetics, pharmaceuticals, and lubricants.

Enterococcus faecalis Readily Adapts Membrane Phospholipid Composition to Environmental and Genetic Perturbation.

The bacterial lipid membrane, consisting both of fatty acid (acyl) tails and polar head groups, responds to changing conditions through alteration of either the acyl tails and/or head groups. This plasticity is critical for cell survival as it allows maintenance of both the protective nature of the membrane as well as functioning membrane protein complexes. Bacteria that live in fatty-acid rich environments, such as those found in the human host, can exploit host fatty acids to synthesize their own membranes, in turn, altering their physiology. Enterococcus faecalis is such an organism: it is a commensal of the mammalian intestine where it is exposed to fatty-acid rich bile, as well as a major cause of hospital infections during which it is exposed to fatty acid containing-serum. Within, we employed an untargeted approach to detect the most common phospholipid species of E. faecalis OG1RF via ultra-high performance liquid chromatography high-resolution mass spectrometry (UHPLC-HRMS). We examined not only how the composition responds upon exposure to host fatty acids but also how deletion of genes predicted to synthesize major polar head groups impact lipid composition. Regardless of genetic background and differing basal lipid composition, all strains were able to alter their lipid composition upon exposure to individual host fatty acids. Specific gene deletion strains, however, had altered survival to membrane damaging agents. Combined, the enterococcal lipidome is highly resilient in response to both genetic and environmental perturbation, likely contributing to stress survival.

How Can Immuno suppression After Organ Transplantation be Made More Effective and Safer? – A Review on Liposomal Formulations with Consideration of Archaeal Tetraetherlipid

Immune-suppressive agents such as methylprednisolone and cyclosporine exert tremendous side effects, because of high dosage and long-term application required for immune suppression after organ transplantation. Major side effects of methylprednisolone include bleeding of the gastro-intestinal tract, hypertension, and osteoporosis, whereas cyclosporine is nephrotoxic. Liposomes are phospholipid particles that allow delivery of drugs preferentially to the reticuloendothelial system. They can be prepared from phospholipids such as lecithin from soybean or egg yolk, other specific or modified lipids or from membrane-spanning tetraether lipid (TEL), which can be extracted and purified from archaeal cell membranes. One advantage in the use of liposomal application is reduced toxicity of many drugs. We report on various liposomal preparations of cyclosporine, methylprednisolone (L-MPL) and its palmitate derivative (L-MPLP). It has been documented that liposomal cyclosporine A (L-CsA), 1.75 mg/kg/day for seven days has potential for use as an immune-suppressive agent in rats with increased efficacy and decreased nephrotoxicity compared to commercially available forms of intravenous CsA. Liposomal methylprednisolone (L-MPL) 2 mg/kg, intravenously (IV), twice a week shows significantly prolonged cardiac allograft survival in rats and tissue-selective sequestration of the drug in comparison with the same dosage regimen of methylprednisolone in solution, administered daily. We report on organ distribution of L-MPLP in rats after intraperitoneal (IP) administration. Conclusion: Liposomal preparations of immunosuppressants have significantly higher immune-suppressive potential and lower toxicity than non-liposomal preparations. Bipolar TEL can be extracted, fractionated and purified from archaea to form stable liposomes which are extremely resistant, even to gastric fluid. Hence, TEL liposomes allow (besides IV and IP) for oral administration of immuno suppressants after organ transplantation with pharmacological and toxicological advantages as common liposomal phospholipid bilayer preparations.

Interfacial Dynamics in Lipid Membranes: The Effects of Headgroup Structures.

Phospholipid membranes support essential biochemical processes, yet remain difficult to characterize due to their compositional and structural heterogeneity. The two most common phospholipid headgroup structures in biological membranes are phosphatidylcholine (PC) and phosphatidylethanolamine (PE), but interactions between PC and PE lipids remain underexplored. In this study, we apply ultrafast two-dimensional infrared (2D IR) spectroscopy to quantify the headgroup effects on interfacial dynamics in PC/PE lipid mixtures. Experiments are interpreted through molecular dynamics simulations using the molecular dynamics with alchemical step (MDAS) algorithm for enhanced sampling. Experimental results indicate that the PE content decreases H-bond formation at the ester carbonyl positions near the lipid membrane's hydrophobic core as a result of increased packing density. The observed dehydration is linked to faster molecular dynamics within the interfacial region.

Interactions of l-arginine with Langmuir monolayers of common phospholipids at the air-water interface

The interactions of l-arginine (l-arg) with Langmuir monolayers of three most common phospholipids, which are sodium salt of dipalmitoylphosphatidylglycerol (DPPG), dipalmitoylphosphatidylcholine (DPPC) and dipalmitoylphosphatidylethanolamine (DPPE), have been investigated at the air-water interface. The surface pressure-area (π-A) isotherms of these monolayers have been measured with a film balance and monolayer morphology has been observed by a Brewster angle microscopy (BAM). The DPPG monolayers on pure water do not show any phase transition but show irregular shaped condensed phases formed just after evaporation of the solvent at 20 °C. However, this monolayer on l-arg solution subphase indicates a first-order phase transition from liquid expanded to liquid condensed (LE-LC) phases and forms LC domains at the same temperature. With an increase in the l-arg concentration in the subphase up to 5.0 × 10−4 M, the π-A shows an overall increasingly greater expansion in the molecular area. All of the π-A isotherms recorded on ≥5.0 × 10−4 M l-arg solution subphases almost coincide with each other. These changes in the phase behavior have been explained by the fact that l-arg having guanidinium cationic group undergoes strong hydrogen bonding interaction with the anionic phosphatidylglycerol (PG-) head group. The bonding between two molecules is further strengthened by electrostatic attraction between cationic l-arg and anionic PG- ions. The BAM observation of the monolayer morphology supports this explanation. On the other hand, a very negligible interaction has been observed between l-arg and DPPC or DPPE monolayers. The π-A isotherms in the presence of l-arg for both the amphiphiles show a very little expansion only in the LE phase region, but coincide in the so called solid phase region. The monolayer morphology of both the monolayers also supports these results. This little effect of expansion in the LE region may be explained by the ion-pair formation between cationic l-arg and anionic head groups in the monolayers at lower pressures. However, due to compression at high pressure, the l-arg molecules are squeezed out from the amphiphile head groups.

Lipids in xylem sap of woody plants across the angiosperm phylogeny.

Lipids have been observed attached to lumen-facing surfaces of mature xylem conduits of several plant species, but there has been little research on their functions or effects on water transport, and only one lipidomic study of the xylem apoplast. Therefore, we conducted lipidomic analyses of xylem sap from woody stems of seven plants representing six major angiosperm clades, including basal magnoliids, monocots and eudicots, to characterize and quantify phospholipids, galactolipids and sulfolipids in sap using mass spectrometry. Locations of lipids in vessels of Laurus nobilis were imaged using transmission electron microscopy and confocal microscopy. Xylem sap contained the galactolipids di- and monogalactosyldiacylglycerol, as well as all common plant phospholipids, but only traces of sulfolipids, with total lipid concentrations in extracted sap ranging from 0.18 to 0.63nmolml-1 across all seven species. Contamination of extracted sap from lipids in cut living cells was found to be negligible. Lipid composition of sap was compared with wood in two species and was largely similar, suggesting that sap lipids, including galactolipids, originate from cell content of living vessels. Seasonal changes in lipid composition of sap were observed for one species. Lipid layers coated all lumen-facing vessel surfaces of L. nobilis, and lipids were highly concentrated in inter-vessel pits. The findings suggest that apoplastic, amphiphilic xylem lipids are a universal feature of angiosperms. The findings require a reinterpretation of the cohesion-tension theory of water transport to account for the effects of apoplastic lipids on dynamic surface tension and hydraulic conductance in xylem.

Characterization of Protein-Phospholipid/Membrane Interactions Using a "Membrane-on-a-Chip" Microfluidic System.

It is now clear that organelles of a mammalian cell can be distinguished by phospholipid profiles, both as ratios of common phospholipids and by the absence or presence of certain phospholipids. Organelle-specific phospholipids can be used to provide a specific shape and fluidity to the membrane and/or used to recruit and/or traffic proteins to the appropriate subcellular location and to restrict protein function to this location. Studying the interactions of proteins with specific phospholipids using soluble derivatives in isolation does not always provide useful information because the context in which the headgroups are presented almost always matters. Our laboratory has shown this circumstance to be the case for a viral protein binding to phosphoinositides in solution and in membranes. The system we have developed to study protein-phospholipid interactions in the context of a membrane benefits from the creation of tailored membranes in a channel of a microfluidic device, with a fluorescent lipid in the membrane serving as an indirect reporter of protein binding. This system is amenable to the study of myriad interactions occurring at a membrane surface as long as a net change in surface charge occurs in response to the binding event of interest.

Protein Sieving with Capillary Nanogel Electrophoresis.

Protein sieving, which is a fundamental tool in the biotechnology field, can be automated using capillary gel electrophoresis. The high-viscosity and biocompatible linear gels required for capillary sieving must be replaced for each run using high pressures. Thermally responsive gels are easier to renew in the capillary as they can be repetitively switched between low- and high-viscosity solutions. A thermally responsive sieving gel was recently demonstrated to separate DNA, which is a larger biomolecule than proteins. This material required no synthesis as it was self-assembled from common phospholipids. Nanogels composed of dimyristoyl-sn-glycero-2-phosphocholine and 1,2-dihexanoyl-sn-glycero-3-phosphocholine exhibit thermally reversible viscosity within a 10 °C temperature change, forming a sieving matrix above 24 °C. Additionally, these nanogels are nondenaturing and have been demonstrated to preserve the activity of enzymes. In this report, a phospholipid nanogel is used for the first time for capillary gel electrophoresis separations of proteins. The mobilities in buffer and nanogel demonstrated that 20-30% nanogel supports sieving of proteins ranging from 20 to 80 kDa. Capillary separations based on sieving rather than electrophoresis had similar precision in both area and migration time as well as similar separation efficiencies. However, the migration time increased with gel concentration. The nanogel was used for the analysis of proteins in human serum. Proteins in the sample were more effectively resolved and quantified with capillary sieving compared to free-solution capillary electrophoresis. This allowed for accurate quantification.

Integration of LC/MS-based molecular networking and classical phytochemical approach allows in-depth annotation of the metabolome of non-model organisms - The case study of the brown seaweed Taonia atomaria

Untargeted LC-MS based metabolomics is a useful approach in many research areas such as medicine, systems biology, environmental sciences or even ecology. In such an approach, annotation of metabolomes of non-model organisms remains a significant challenge. In this study, an analytical workflow combining a classical phytochemical approach, using the isolation and the full characterization of the chemical structure of natural products, together with the use of MS/MS-based molecular networking with various levels of restrictiveness was developed. This protocol was applied to the marine brown seaweed Taonia atomaria, a cosmopolitan algal species, and allowed to annotate more than 200 metabolites. First, the algal organic crude extracts were fractionated by flash-chromatography and the chemical structure of eight of the main chemical constituents of this alga were fully characterized by means of spectroscopic methods (1D and 2D NMR, HRMS). These compounds were further used as chemical standards. In a second step, the main fractions of the algal extracts were analyzed by UHPLC-MS/MS and the resulting data were uploaded to the Global Natural Products Social Molecular Networking platform (GNPS) to create several molecular networks (MNs). A first MN (MN-1) was built with restrictive parameters and allowed the creation of clusters composed by nodes with highly similar MS/MS spectra. Then, using database hits and chemical standards as “seed” nodes and/or similarity between MS/MS fragmentation pattern, the main clusters were easily annotated as common glycerolipids and phospholipids, much rare lipids -such as acylglycerylhydroxymethyl-N,N,N-trimethyl-ß-alanines or fulvellic acid derivatives- but also new glycerolipids bearing a terpene moiety. Lastly, the use of less and less constrained MNs allowed to further increase the number of annotated metabolites.

Polar Head Modified Phospholipids by Phospholipase D-Catalyzed Transformations of Natural Phosphatidylcholine for Targeted Applications: An Overview

This review describes the use of phospholipase D (PLD) to perform the transphosphatidylation of the most common natural phospholipid (PL), phosphatidylcholine (PC) to obtain polar head modified phospholipids with real targeted applications. The introduction of different polar heads with distinctive physical and chemical properties such as charge, polarity and dimensions allows the obtainment of very different PLs, which can be exploited in very diverse fields of application. Moreover, the inclusions of a bioactive moiety in the PL polar head constitutes a powerful tool for the stabilization and administration of active ingredients. The use of this biocatalytic approach allows the preparation of compounds which cannot be easily obtained by classical chemical methods, by using mild and green reaction conditions. PLD is a very versatile enzyme, able to catalyze both the hydrolysis of PC to choline and phosphatidic acid (PA), and the transphosphatidylation reaction in the presence of an appropriate alcohol. The yield of production of the desired product and the ratio with the collateral PA formation is highly dependent on parameters such as the nature and concentration of the alcohol and the enzymatic source. The application of PLD catalyzed transformations for the production of a great number of PLs with important uses in medical, nutraceutical and cosmetic sectors will be discussed in this work.

Interactions of cryoprotective agents with phospholipid membranes - A Langmuir monolayer study

The influence of four common cryoprotectants (dimethyl sulfoxide, glycerol, ethylene glycol and dimethylformamide) on monolayers of four common phospholipids (DPPC, DOPC, POPC and POPE) have been studied using Langmuir isotherms and monolayer insertion experiments. The cryoprotectant concentrations were chosen to be directly relevant to cryoprotection. We show that DMSO causes an expansion of the DPPC area per lipid (in contrast to previous work at higher concentrations). However, it caused compression for POPC, and had little effect for POPE or DOPC. As most previous studies have involved only DPPC, this highlights the importance of studying different lipid types as these may have a significant effect on the interactions. We show that both ethylene glycol and glycerol cause a small expansion of the monolayer at fixed pressure, implying that they insert into the headgroup regions, regardless of lipid species, and consistent with their ability to penetrate membranes. By contrast, dimethylformamide causes monolayer compression for all lipid species, implying it dehydrates the lipid head groups. Membrane insertion experiments at physiological values of lateral pressure highlight that DPPC is the most difficult lipid to penetrate, implying that the penetrating action of cryoprotectants may only occur for unsaturated phospholipids. Thus, extrapolations of results based solely on the DPPC need to be made with care.

Gas-Phase Protonation Thermodynamics of Biological Lipids: Experiment, Theory, and Implications.

Phospholipids are important to cellular function and are a vital structural component of plasma and organelle membranes. These membranes isolate the cell from its environment, allow regulation of the internal concentrations of ions and small molecules, and host diverse types of membrane proteins. It remains extremely challenging to identify specific membrane protein-lipid interactions and their relative strengths. Native mass spectrometry, an intrinsically gas-phase method, has recently been demonstrated as a promising tool for identifying endogenous protein-lipid interactions. However, to what extent the identified interactions reflect solution- versus gas-phase binding strengths is not known. Here, the "Extended" Kinetic Method and ab initio computations at three different levels of theory are used to experimentally and theoretically determine intrinsic gas-phase basicities (GB, ΔG for deprotonation of the protonated base) and proton affinities (PA, ΔH for deprotonation of the protonated base) of six lipids representing common phospholipid types. Gas-phase acidities (ΔG and ΔH for deprotonation) of neutral phospholipids are also evaluated computationally and ranked experimentally. Intriguingly, it is found that two of these phospholipids, sphingomyelin and phosphatidylcholine, have the highest GB of any small, monomeric biomolecules measured to date and are more basic than arginine. Phosphatidylethanolamine and phosphatidylserine are found to be similar in GB to basic amino acids lysine and histidine, and phosphatidic acid and phosphatidylglycerol are the least basic of the six lipid types studied, though still more basic than alanine. Kinetic Method experiments and theory show that the gas-phase acidities of these phospholipids are high but less extreme than their GB values, with phosphatidylserine and phosphatidylglycerol being the most acidic. These results indicate that sphingomyelin and phosphatidylcholine lipids can act as charge-reducing agents when dissociated from native membrane protein-lipid complexes in the gas phase and provide a straightforward model to explain the results of several recent native mass spectrometry studies of protein-lipid complexes.

Identifying lipids tightly bound to an integral membrane protein

Anabaena Sensory Rhodopsin (ASR) is a microbial photosensor from the cyanobacterium Anabaena sp. PCC 7120. It was found in previous studies that ASR co-purifies with several small molecules, although their identities and structural or functional roles remained unclear. Here, we use solid-state nuclear magnetic resonance (SSNMR) spectroscopy and mass spectrometry to characterize these molecules. Numerous correlations atypical for protein amino acids were found and assigned in the SSNMR spectra. The chemical shift patterns correspond to N-acetyl-d-glucosamine, N-acetyl-d-mannosaminuronic acid, and 4-acetamido-4,6-dideoxy-d-galactose which are part of the Enterobacterial Common Antigen (ECA). These sugars undergo rapid anisotropic motions and are likely linked flexibly to a rigid anchor that tightly binds ASR. Phosphorus NMR reveals several signals that are characteristic of monophosphates, further suggesting phosphatidylglyceride as the ECA lipid carrier which is anchored to ASR. In addition, NMR signals corresponding to common phospholipid phosphatidylethanolamine (PE) have been detected. The presence of PE tightly interacting with ASR was confirmed using liquid chromatography-mass spectrometry. This article commemorates Professor Michèle Auger and her contributions to membrane biophysics and Nuclear Magnetic Resonance.

Common disbalance in the brain parenchyma of dementias: Phospholipid profile analysis between CADASIL and sporadic Alzheimer's disease

Sporadic Alzheimer's disease (SAD) is the most common form of dementia, and cerebral autosomal dominant arteriopathy with subcortical infarcts and leukoencephalopathy (CADASIL) is the most frequent hereditary ischemic small vessel disease of the brain. Relevant biomarkers or specific metabolic signatures could provide powerful tools to manage these diseases. Therefore, the main goal of this study was to compare the postmortem frontal cortex gray matter, white matter and cerebrospinal fluid (CSF) between a cognitively healthy group and CADASIL and SAD groups. We evaluated 352 individual lipids, belonging to 13 lipid classes/subclasses, using mass spectrometry, and the lipid profiles were subjected to multivariate analysis to discriminate between the dementia groups (CADASIL and SAD) and healthy controls. The main lipid molecular species showing greater discrimination by partial least squares-discriminant analysis (PLS-DA) and a higher significance multivariate correlation (sMC) index were as follows: phosphatidylserine (PS) PS(44:7) and lysophosphatidylethanolamine (LPE) LPE(18:2) in gray matter (GM); phosphatidylethanolamine (PE) PE(32:2) and phosphatidylcholine PC PC(44:6) in white matter (WM), and ether PE (ePE) ePE(38:2) and ether PC (ePC) ePC(34:3) in CSF. Common phospholipid molecular species were obtained in both dementias, such as PS(44:7) and lyso PC (LPC) LPC(22:5) in GM, PE(32:2) in WM and phosphatidic acid (PA) PA(38:5) and PC(42:7) in CFS. Our exploratory study suggests that phospholipids (PLs) involved in neurotransmission alteration, connectivity impairment and inflammation response in GM, WM and CSF are a transversal phenomenon affecting dementias such as CADASIL and SAD independent of the etiopathogenesis, thus providing a possible common prodromal phospholipidic biomarker of dementia.