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Nature Feb 2024The outer membrane in Gram-negative bacteria consists of an asymmetric phospholipid-lipopolysaccharide bilayer that is densely packed with outer-membrane β-barrel...
The outer membrane in Gram-negative bacteria consists of an asymmetric phospholipid-lipopolysaccharide bilayer that is densely packed with outer-membrane β-barrel proteins (OMPs) and lipoproteins. The architecture and composition of this bilayer is closely monitored and is essential to cell integrity and survival. Here we find that SlyB, a lipoprotein in the PhoPQ stress regulon, forms stable stress-induced complexes with the outer-membrane proteome. SlyB comprises a 10 kDa periplasmic β-sandwich domain and a glycine zipper domain that forms a transmembrane α-helical hairpin with discrete phospholipid- and lipopolysaccharide-binding sites. After loss in lipid asymmetry, SlyB oligomerizes into ring-shaped transmembrane complexes that encapsulate β-barrel proteins into lipid nanodomains of variable size. We find that the formation of SlyB nanodomains is essential during lipopolysaccharide destabilization by antimicrobial peptides or acute cation shortage, conditions that result in a loss of OMPs and compromised outer-membrane barrier function in the absence of a functional SlyB. Our data reveal that SlyB is a compartmentalizing transmembrane guard protein that is involved in cell-envelope proteostasis and integrity, and suggest that SlyB represents a larger family of broadly conserved lipoproteins with 2TM glycine zipper domains with the ability to form lipid nanodomains.
Topics: Bacterial Outer Membrane Proteins; Cell Membrane; Glycine; Lipopolysaccharides; Lipoproteins; Phospholipids; Binding Sites; Proteostasis; Lipid Bilayers; Proteome; Regulon; Protein Domains; Antimicrobial Peptides; Gram-Negative Bacteria
PubMed: 38081298
DOI: 10.1038/s41586-023-06925-5 -
ACS Chemical Neuroscience Jun 2021Cooperative binding is a key feature of metabolic pathways, signaling, and transport processes. It provides tight regulation over a narrow concentration interval of a...
Cooperative binding is a key feature of metabolic pathways, signaling, and transport processes. It provides tight regulation over a narrow concentration interval of a ligand, thus enabling switching to be triggered by small concentration variations. The data presented in this work reveal strong positive cooperativity of α-synuclein binding to phospholipid membranes. Fluorescence cross-correlation spectroscopy, confocal microscopy, and -TEM results show that in excess of vesicles α-synuclein does not distribute randomly but binds only to a fraction of all available vesicles. Furthermore, α-synuclein binding to a supported lipid bilayer observed with total internal reflection fluorescence microscopy displays a much steeper dependence of bound protein on total protein concentration than expected for independent binding. The same phenomenon was observed in the case of α-synuclein binding to unilamellar vesicles of sizes in the nm and μm range as well as to flat supported lipid bilayers, ruling out that nonuniform binding of the protein is governed by differences in membrane curvature. Positive cooperativity of α-synuclein binding to lipid membranes means that the affinity of the protein to a membrane is higher where there is already protein bound compared to a bare membrane. The phenomenon described in this work may have implications for α-synuclein function in synaptic transmission and other membrane remodeling events.
Topics: Lipid Bilayers; Phospholipids; Protein Binding; Spectrometry, Fluorescence; Unilamellar Liposomes; alpha-Synuclein
PubMed: 34076426
DOI: 10.1021/acschemneuro.1c00006 -
International Journal of Biological... Jul 2022Autophagy is a process in which parts of the eukaryotic cell are selectively degraded in the lysosome. The materials to be catabolized are first surrounded by a...
Autophagy is a process in which parts of the eukaryotic cell are selectively degraded in the lysosome. The materials to be catabolized are first surrounded by a double-membrane structure, the autophagosome. Autophagosome generation is a complex event, in which many proteins are involved. Among the latter, yeast Atg8 or its mammalian orthologues are essential in autophagosome membrane elongation, shaping and closure. A subfamily of the human Atg8 orthologues is formed by the proteins LC3A, LC3B, and LC3C. Previous studies suggest that, at variance with the other two, LC3C does not participate in cardiolipin-mediated mitophagy. The present study was devoted to exploring the binding of LC3C to lipid vesicles, bilayers and monolayers, and the ensuing protein-dependent perturbing effects, in the absence of the mitochondrial lipid cardiolipin. All Atg8 orthologues are covalently bound to a phospholipid prior to their involvement in autophagosome elongation. In our case, a mutant in the C-terminal amino acid, LC3C G126C, together with the use of a maleimide-derivatized phosphatidyl ethanolamine, ensured LC3C lipidation, up to 100% under certain conditions. Ultracentrifugation, surface pressure measurements, spectroscopic and cryo-electron microscopic techniques revealed that lipidated LC3C induced vesicle aggregation (5-fold faster in sonicated than in large unilamellar vesicles) and inter-vesicular lipid mixing (up to 82%), including inner-monolayer lipid mixing (up to 32%), consistent with in vitro partial vesicle fusion. LC3C was also able to cause the release of 80-90% vesicular aqueous contents. The data support the idea that LC3C would be able to help in autophagosome elongation/fusion in autophagy phenomena.
Topics: Autophagy; Cardiolipins; Humans; Microtubule-Associated Proteins; Phospholipids; Protein Binding; Saccharomyces cerevisiae
PubMed: 35618088
DOI: 10.1016/j.ijbiomac.2022.05.129 -
Life Science Alliance Feb 2023Autotaxin is primarily known for the formation of lysophosphatidic acid (LPA) from lysophosphatidylcholine. LPA is an important signaling phospholipid that can bind to...
Autotaxin is primarily known for the formation of lysophosphatidic acid (LPA) from lysophosphatidylcholine. LPA is an important signaling phospholipid that can bind to six G protein-coupled receptors (LPA). The ATX-LPA signaling axis is a critical component in many physiological and pathophysiological conditions. Here, we describe a potent inhibition of Δ--tetrahydrocannabinol (THC), the main psychoactive compound of medicinal cannabis and related cannabinoids, on the catalysis of two isoforms of ATX with nanomolar apparent EC values. Furthermore, we decipher the binding interface of ATX to THC, and its derivative 9(R)-Δ6a,10a-THC (6a10aTHC), by X-ray crystallography. Cellular experiments confirm this inhibitory effect, revealing a significant reduction of internalized LPA in the presence of THC with simultaneous ATX and lysophosphatidylcholine stimulation. Our results establish a functional interaction of THC with autotaxin-LPA signaling and highlight novel aspects of medicinal cannabis therapy.
Topics: Lysophosphatidylcholines; Lysophospholipids; Medical Marijuana; Receptors, Lysophosphatidic Acid; Dronabinol
PubMed: 36623871
DOI: 10.26508/lsa.202201595 -
Biomolecules Jun 2023G protein-coupled receptors (GPCRs) are embedded in phospholipid membrane bilayers with cholesterol representing 34% of the total lipid content in mammalian plasma... (Review)
Review
G protein-coupled receptors (GPCRs) are embedded in phospholipid membrane bilayers with cholesterol representing 34% of the total lipid content in mammalian plasma membranes. Membrane lipids interact with GPCRs structures and modulate their function and drug-stimulated signaling through conformational selection. It has been shown that anionic phospholipids form strong interactions between positively charged residues in the G protein and the TM5-TM6-TM 7 cytoplasmic interface of class A GPCRs stabilizing the signaling GPCR-G complex. Cholesterol with a high content in plasma membranes can be identified in more specific sites in the transmembrane region of GPCRs, such as the Cholesterol Consensus Motif (CCM) and Cholesterol Recognition Amino Acid Consensus (CRAC) motifs and other receptor dependent and receptor state dependent sites. Experimental biophysical methods, atomistic (AA) MD simulations and coarse-grained (CG) molecular dynamics simulations have been applied to investigate these interactions. We emphasized here the impact of phosphatidyl inositol-4,5-bisphosphate (PtdIns(4,5)P or PIP), a minor phospholipid component and of cholesterol on the function-related conformational equilibria of the human A adenosine receptor (AR), a representative receptor in class A GPCR. Several GPCRs of class A interacted with PIP and cholesterol and in many cases the mechanism of the modulation of their function remains unknown. This review provides a helpful comprehensive overview for biophysics that enter the field of GPCRs-lipid systems.
Topics: Animals; Humans; Binding Sites; Cholesterol; Mammals; Molecular Dynamics Simulation; Phospholipids; Receptors, G-Protein-Coupled; Receptors, Purinergic P1; Receptor, Adenosine A2A
PubMed: 37371538
DOI: 10.3390/biom13060957 -
The Journal of Biological Chemistry Jun 2023The human AdipoR2 and its Caenorhabditis elegans homolog PAQR-2 are multipass plasma membrane proteins that protect cells against membrane rigidification. However, how...
The human AdipoR2 and its Caenorhabditis elegans homolog PAQR-2 are multipass plasma membrane proteins that protect cells against membrane rigidification. However, how AdipoR2 promotes membrane fluidity mechanistically is not clear. Using 13C-labeled fatty acids, we show that AdipoR2 can promote the elongation and incorporation of membrane-fluidizing polyunsaturated fatty acids into phospholipids. To elucidate the molecular basis of these activities, we performed immunoprecipitations of tagged AdipoR2 and PAQR-2 expressed in HEK293 cells or whole C. elegans, respectively, and identified coimmunoprecipitated proteins using mass spectrometry. We found that several of the evolutionarily conserved AdipoR2/PAQR-2 interactors are important for fatty acid elongation and incorporation into phospholipids. We experimentally verified some of these interactions, namely, with the dehydratase HACD3 that is essential for the third of four steps in long-chain fatty acid elongation and ACSL4 that is important for activation of unsaturated fatty acids and their channeling into phospholipids. We conclude that AdipoR2 and PAQR-2 can recruit protein interactors to promote the production and incorporation of unsaturated fatty acids into phospholipids.
Topics: Animals; Humans; Caenorhabditis elegans; Caenorhabditis elegans Proteins; Cell Membrane; Fatty Acids; HEK293 Cells; Membrane Fluidity; Phospholipids; Receptors, Adiponectin; Protein Binding
PubMed: 37164154
DOI: 10.1016/j.jbc.2023.104799 -
Chemphyschem : a European Journal of... Aug 2020Ions are ubiquitous in nature. They play a key role for many biological processes on the molecular scale, from molecular interactions, to mechanical properties, to... (Review)
Review
Ions are ubiquitous in nature. They play a key role for many biological processes on the molecular scale, from molecular interactions, to mechanical properties, to folding, to self-organisation and assembly, to reaction equilibria, to signalling, to energy and material transport, to recognition etc. Going beyond monovalent ions to multivalent ions, the effects of the ions are frequently not only stronger (due to the obviously higher charge), but qualitatively different. A typical example is the process of binding of multivalent ions, such as Ca , to a macromolecule and the consequences of this ion binding such as compaction, collapse, potential charge inversion and precipitation of the macromolecule. Here we review these effects and phenomena induced by multivalent ions for biological (macro)molecules, from the "atomistic/molecular" local picture of (potentially specific) interactions to the more global picture of phase behaviour including, e. g., crystallisation, phase separation, oligomerisation etc. Rather than attempting an encyclopedic list of systems, we rather aim for an embracing discussion using typical case studies. We try to cover predominantly three main classes: proteins, nucleic acids, and amphiphilic molecules including interface effects. We do not cover in detail, but make some comparisons to, ion channels, colloidal systems, and synthetic polymers. While there are obvious differences in the behaviour of, and the relevance of multivalent ions for, the three main classes of systems, we also point out analogies. Our attempt of a comprehensive discussion is guided by the idea that there are not only important differences and specific phenomena with regard to the effects of multivalent ions on the main systems, but also important similarities. We hope to bridge physico-chemical mechanisms, concepts of soft matter, and biological observations and connect the different communities further.
Topics: Cations; Lipid Bilayers; Metals; Micelles; Nucleic Acids; Phospholipids; Protein Binding; Proteins; Static Electricity
PubMed: 32406605
DOI: 10.1002/cphc.202000162 -
Nutrients May 2022L-serine plays an essential role in a broad range of cellular functions including protein synthesis, neurotransmission, and folate and methionine cycles and synthesis of... (Review)
Review
L-serine plays an essential role in a broad range of cellular functions including protein synthesis, neurotransmission, and folate and methionine cycles and synthesis of sphingolipids, phospholipids, and sulphur containing amino acids. A hydroxyl side-chain of L-serine contributes to polarity of proteins, and serves as a primary site for binding a phosphate group to regulate protein function. D-serine, its D-isoform, has a unique role. Recent studies indicate increased requirements for L-serine and its potential therapeutic use in some diseases. L-serine deficiency is associated with impaired function of the nervous system, primarily due to abnormal metabolism of phospholipids and sphingolipids, particularly increased synthesis of deoxysphingolipids. Therapeutic benefits of L-serine have been reported in primary disorders of serine metabolism, diabetic neuropathy, hyperhomocysteinemia, and amyotrophic lateral sclerosis. Use of L-serine and its metabolic products, specifically D-serine and phosphatidylserine, has been investigated for the therapy of renal diseases, central nervous system injury, and in a wide range of neurological and psychiatric disorders. It is concluded that there are disorders in which humans cannot synthesize L-serine in sufficient quantities, that L-serine is effective in therapy of disorders associated with its deficiency, and that L-serine should be classified as a "conditionally essential" amino acid.
Topics: Amino Acid Metabolism, Inborn Errors; Amino Acids, Essential; Humans; Phospholipids; Serine; Sphingolipids
PubMed: 35565953
DOI: 10.3390/nu14091987 -
International Journal of Molecular... Sep 2023Several discoveries show that with age and cataract formation, β-crystallin binds with the lens membrane or associates with other lens proteins, which bind with the...
Several discoveries show that with age and cataract formation, β-crystallin binds with the lens membrane or associates with other lens proteins, which bind with the fiber cell plasma membrane, accompanied by light scattering and cataract formation. However, how lipids (phospholipids and sphingolipids) and cholesterol (Chol) influence β-crystallin binding to the membrane is unclear. This research aims to elucidate the role of lipids and Chol in the binding of β-crystallin to the membrane and the membrane's physical properties (mobility, order, and hydrophobicity) with β-crystallin binding. We used electron paramagnetic resonance (EPR) spin-labeling methods to investigate the binding of β-crystallin with a model of porcine lens-lipid (MPLL), model of mouse lens-lipid (MMLL), and model of human lens-lipid (MHLL) membrane with and without Chol. Our results show that β-crystallin binds with all of the investigated membranes in a saturation manner, and the maximum parentage of the membrane surface occupied (MMSO) by β-crystallin and the binding affinity (K) of β-crystallin to the membranes followed trends: MMSO (MPLL) > MMSO (MMLL) > MMSO (MHLL) and K (MHLL) > K (MMLL) ≈ K (MPLL), respectively, in which the presence of Chol reduces the MMSO and K for all membranes. The mobility near the headgroup regions of the membranes decreases with an increase in the binding of β-crystallin; however, the decrease is more pronounced in the MPLL and MMLL membranes than the MHLL membrane. In the MPLL and MMLL membranes, the membranes become slightly ordered near the headgroup with an increase in β-crystallin binding compared to the MHLL membrane. The hydrophobicity near the headgroup region of the membrane increases with β-crystallin binding; however, the increase is more pronounced in the MPLL and MMLL membranes than the MHLL membrane, indicating that β-crystallin binding creates a hydrophobic barrier for the passage of polar molecules, which supports the barrier hypothesis in cataract formation. However, in the presence of Chol, there is no significant increase in hydrophobicity with β-crystallin binding, suggesting that Chol prevents the formation of a hydrophobic barrier, possibly protecting against cataract formation.
Topics: Mice; Humans; Animals; Swine; Crystallins; Lens, Crystalline; beta-Crystallins; Cataract; Phospholipids
PubMed: 37686406
DOI: 10.3390/ijms241713600 -
Frontiers in Endocrinology 2020A huge diversification of phospholipids, forming the aqueous interfaces of all biomembranes, cannot be accommodated within a simple concept of their role as membrane... (Review)
Review
A huge diversification of phospholipids, forming the aqueous interfaces of all biomembranes, cannot be accommodated within a simple concept of their role as membrane building blocks. Indeed, a number of signaling functions of (phospho)lipid molecules has been discovered. Among these signaling lipids, a particular group of oxygenated polyunsaturated fatty acids (PUFA), so called lipid mediators, has been thoroughly investigated over several decades. This group includes oxygenated octadecanoids, eicosanoids, and docosanoids and includes several hundreds of individual species. Oxygenation of PUFA can occur when they are esterified into major classes of phospholipids. Initially, these events have been associated with non-specific oxidative injury of biomembranes. An alternative concept is that these post-synthetically oxidatively modified phospholipids and their adducts with proteins are a part of a redox epiphospholipidome that represents a rich and versatile language for intra- and inter-cellular communications. The redox epiphospholipidome may include hundreds of thousands of individual molecular species acting as meaningful biological signals. This review describes the signaling role of oxygenated phospholipids in programs of regulated cell death. Although phospholipid peroxidation has been associated with almost all known cell death programs, we chose to discuss enzymatic pathways activated during apoptosis and ferroptosis and leading to peroxidation of two phospholipid classes, cardiolipins (CLs) and phosphatidylethanolamines (PEs). This is based on the available LC-MS identification and quantitative information on the respective peroxidation products of CLs and PEs. We focused on molecular mechanisms through which two proteins, a mitochondrial hemoprotein cytochrome (cyt ), and non-heme Fe lipoxygenase (LOX), change their catalytic properties to fulfill new functions of generating oxygenated CL and PE species. Given the high selectivity and specificity of CL and PE peroxidation we argue that enzymatic reactions catalyzed by cyt /CL complexes and 15-lipoxygenase/phosphatidylethanolamine binding protein 1 (15LOX/PEBP1) complexes dominate, at least during the initiation stage of peroxidation, in apoptosis and ferroptosis. We contrast cell-autonomous nature of CLox signaling in apoptosis correlating with its anti-inflammatory functions non-cell-autonomous ferroptotic signaling facilitating pro-inflammatory (necro-inflammatory) responses. Finally, we propose that small molecule mechanism-based regulators of enzymatic phospholipid peroxidation may lead to highly specific anti-apoptotic and anti-ferroptotic therapeutic modalities.
Topics: Animals; Apoptosis; Catalysis; Cell Death; Fatty Acids, Unsaturated; Ferroptosis; Humans; Lipidomics; Oxidation-Reduction; Phospholipids; Signal Transduction
PubMed: 33679610
DOI: 10.3389/fendo.2020.628079