Probing the Membrane Deformations Induced by Binding of Membrane Proteins: Alpha-Synuclein and CRAC (original) (raw)
Related papers
Biophysical Journal, 2011
PMP1, a regulatory subunit of the yeast plasma membrane H þ -ATPase, is a single transmembrane helix protein. Its cytoplasmic C-terminus possesses several positively charged residues and interacts with phosphatidylserine lipids as shown through both 1 H-and 2 H-NMR experiments. We used all-atom molecular dynamics simulations to obtain atomic-scale data on the effects of membrane interface lipid composition on PMP1 structure and tilt. PMP1 was embedded in two hydrated bilayers, differing in the composition of the interfacial region. The neutral bilayer is composed of POPC (1-palmitoyl-2-oleoyl-3-glycerophosphatidylcholine) lipids and the negatively charged bilayer is composed of POPC and anionic POPS (1-palmitoyl-2-oleoyl-3glycero-phosphatidylserine) lipids. Our results were consistent with NMR data obtained previously, such as a lipid sn-2 chain lying on the W28 aromatic ring and in the groove formed on one side of the PMP1 helix. In pure POPC, the transmembrane helix is two residues longer than the initial structure and the helix tilt remains constant at 6 5 3 . By contrast, in mixed POPC-POPS, the initial helical structure of PMP1 is stable throughout the simulation time even though the C-terminal residues interact strongly with POPS headgroups, leading to a significant increase of the helix tilt within the membrane to 20 5 5 .
Investigation of the Binding Geometry of a Peripheral Membrane Protein †
Biochemistry, 2005
A growing number of modules including FYVE domains target key signaling proteins to membranes through specific recognition of lipid headgroups and hydrophobic insertion into bilayers. Despite the critical role of membrane insertion in the function of these modules, the structural mechanism of membrane docking and penetration remains unclear. In particular, the three dimensional orientation of the inserted proteins with respect to the membrane surface is difficult to define quantitatively. Here, we determined the geometry of the micelle penetration of early endosome antigen 1 (EEA1) FYVE domain by obtaining NMR-derived restraints that correlate with the distances between protein backbone amides and spin labeled probes. The 5-and 14-doxyl-phosphatidylcholine spin labels were incorporated into dodecylphosphocholine (DPC) micelles, and the reduction of amide signal intensities of the FYVE domain due to paramagnetic relaxation enhancement was measured. The vector of the FYVE domain insertion was estimated relative to the molecular axis by minimizing the paramagnetic restraints obtained in phosphatidylinositol 3-phosphate (PI3P)-enriched micelles containing only DPC or mixed with phosphatidylserine (PS). Additional distance restraints were obtained using a novel spin label mimetic of PI(3)P that contains a nitroxyl radical near the threitol group of the lipid. Conformational changes indicative of elongation of the membrane insertion loop (MIL) were detected upon micelle interaction, in which the hydrophobic residues of the loop tend to move deeper into the non-polar core of micelles. The micelle insertion mechanism of the FYVE domain defined in this study is consistent with mutagenesis data and chemical shift perturbations and demonstrates the advantage of using the spin label NMR approach for investigating the binding geometry by peripheral membrane proteins.
Self-Induced Docking Site of a Deeply Embedded Peripheral Membrane Protein
Biophysical Journal, 2007
As a first step toward understanding the principles of the targeting of C2 domains to membranes, we have carried out a molecular dynamics simulation of the C2 domain of cytosolic phospholipase A2 (cPLA2-C2) in a 1-palmitoyl-2-oleoylphosphatidylcholine bilayer at constant pressure and temperature (NPT, 300 K and 1 atm). Using the high-resolution crystal structure of cPLA2-C2 as a starting point, we embedded two copies of the C2 domain into a preequilibrated membrane at the depth and orientation previously defined by electron paramagnetic resonance (EPR). Noting that in the membrane-bound state the three calcium binding loops are complexed to two calcium ions, we initially restrained the calcium ions at the membrane depth determined by EPR. But the depth and orientation of the domains remained within EPR experimental errors when the restraints were later removed. We find that the thermally disordered, chemically heterogeneous interfacial zones of phosphatidylcholine bilayers allow local lipid remodeling to produce a nearly perfect match to the shape and polarity of the C2 domain, thereby enabling the C2 domain to assemble and optimize its own lipid docking site. The result is a cuplike docking site with a hydrophobic bottom and hydrophilic rim. Contrary to expectations, we did not find direct interactions between the protein-bound calcium ions and lipid headgroups, which were sterically excluded from the calcium binding cleft. Rather, the lipid phosphate groups provided outer-sphere calcium coordination through intervening water molecules. These results show that the combined use of high-resolution protein structures, EPR measurements, and molecular dynamics simulations provides a general approach for analyzing the molecular interactions between membrane-docked proteins and lipid bilayers.
Towards an understanding of membrane-mediated protein–protein interactions
Faraday Discussions, 2010
We propose a computational framework to study the lipid-mediated clustering of integral membrane proteins. Our method employs a hierarchical approach. The potential of mean force (PMF) of two interacting proteins is computed under a coarse-grained 3-D model that successfully describes the structural properties of reconstituted lipid bilayers of dymiristoylphophatidylcholine (DMPC) molecules. Subsequently, a 2-D model is adopted, where proteins represented as self-avoiding disks interact through the previously computed PMF, which is modified to take into account three body corrections. The aggregation of the proteins is extensively studied under the condition of negative hydrophobic mismatch: the formation of clusters with increasing size agrees with previous computational and experimental findings.
Transmembrane domains interactions within the membrane milieu: Principles, advances and challenges
Biochimica et Biophysica Acta (BBA) - Biomembranes, 2012
Protein-protein interactions within the membrane are involved in many vital cellular processes. Consequently, deficient oligomerization is associated with known diseases. The interactions can be partially or fully mediated by transmembrane domains (TMD). However, in contrast to soluble regions, our knowledge of the factors that control oligomerization and recognition between the membrane-embedded domains is very limited. Due to the unique chemical and physical properties of the membrane environment, rules that apply to interactions between soluble segments are not necessarily valid within the membrane. This review summarizes our knowledge on the sequences mediating TMD-TMD interactions which include conserved motifs such as the GxxxG, QxxS, glycine and leucine zippers, and others. The review discusses the specific role of polar, charged and aromatic amino acids in the interface of the interacting TMD helices. Strategies to determine the strength, dynamics and specificities of these interactions by experimental (ToxR, TOXCAT, GALLEX and FRET) or various computational approaches (molecular dynamic simulation and bioinformatics) are summarized. Importantly, the contribution of the membrane environment to the TMD-TMD interaction is also presented. Studies utilizing exogenously added TMD peptides have been shown to influence in vivo the dimerization of intact membrane proteins involved in various diseases. The chirality independent TMD-TMD interactions allows for the design of novel short D-and L-amino acids containing TMD peptides with advanced properties. Overall these studies shed light on the role of specific amino acids in mediating the assembly of the TMDs within the membrane environment and their contribution to protein function. This article is part of a Special Issue entitled: Protein Folding in Membranes.
Membrane Lipid Switches: How Membrane Lipid Structure Influences Protein–Lipid Interactions
2020
Amphitropic membrane proteins are required for signal propagation upon ligand-induced receptor activation at the plasma membrane. These proteins are only activated by ligand-receptor complexes when they both come into physical contact. The interaction between membrane receptors and the amphitropic proteins may not only depend on the expression of these proteins but also on the presence of the peripheral proteins in the vicinity of the membrane receptor, which may be controlled by membrane lipids. Therefore, changes in the membrane lipid composition can induce important changes in cell physiology that affect proliferation, differentiation, and/or cell death. These interactions and the signals they produce are responsible for the pathophysiological status of the cell, which may be influenced by external cues, genetic alterations, lipid storage disorders, etc.. Recent studies have shown that the type and levels of peripheral amphitropic signaling proteins in membranes or aqueous compartments depends on both the membrane's lipid composition, and the protein's amino acid sequence and post-translational lipid modifications. In this sense, alterations in the balance of peripheral signaling proteins at membranes and in the cytosol have been associated with a variety of pathologies. The regulation of membrane lipids controls the type and abundance of the proteins in membranes, an approach that can be used to treat several conditions, including cancer, Alzheimer's disease (AD), cardiovascular diseases (CVDs), inflammation, etc.. This entry aims to review the interaction of amphitropic signaling proteins with membrane structures. This type of interaction deserves further attention because: (i) the plasma membrane is a critical hub for signaling proteins; (ii) cells can regulate their lipid composition according to a range of pathophysiological situations; (iii) membrane lipids organize into different microdomains rich in specific lipid species, which attract different types of proteins; and (iv) proteins that prefer certain types of lipid structures can drive productive interactions involving the reception and propagation of cell signals in certain types of microdomains. In this context, the ability of membranes to generate microdomains is due to the non-homogeneous mixing of membrane lipids. One example of this heterogenous lipid mixture is the transbilayer lipid asymmetry. Higher levels of sphingomyelin (SM) and phosphatidylcholine (PC) have been found in the outer plasma membrane leaflet, whereas phosphatidylethanolamine (PE) and phosphatidylserine (PS) are more abundant in the inner leaflet. This asymmetry has a relevant impact on the biophysical properties of the membrane and the protein-lipid
Transmembrane interactions and the mechanisms of transport of proteins across membranes
Journal of Supramolecular Structure, 1978
We have made observations, by double fluorescence staining of the same cell, of the distributions of surface receptors, and of intracellular actin and myosin, on cultured normal fibroblasts and other flat cells, and on lymphocytes and other rounded cells. The binding of inultivalent ligands (a lectin or specific antibodies) to a cell surface receptor on flat cells clusters the cell receptors into small patches, which line up directly over the actin-and myosin-containing stress fibers inside the cell. Similar ligands binding t o rounded cells can cause their surface receptors t o be collected into caps on the surface, and these caps are invariably found to be associated with concentrations of actin and myosin under the capped membrane. Although these ligand-induced surface phenomena appear to be different o n flat and rounded cells, we propose that in both cases clusters of receptors become linked across the membrane to actin-and myosincontaining structures. In flat cells these structures are very long stress fibers; therefore, when clusters of receptors become linked t o these fibers, the clusters are immobilized. In round cells, membrane-associated actin-and myosincontaining structures are apparently much less extensive than in flat cells; therefore, clusters of receptors linked t o these structures are still mobile in the plane of the membrane. We suggest that in this case the clusters are then actively collected into a cap by an analogue of the muscle sliding filament mechanism.
Journal of the American Chemical Society, 2014
We have investigated the membrane remodeling capacity of the N-terminal membrane-binding domain of α-synuclein (α-Syn 100). Using fluorescence correlation spectroscopy and vesicle clearance assays, we show that α-Syn 100 fully tubulates POPG vesicles, the first demonstration that the amphipathic helix on its own is capable of this effect. We also show that at equal density of membrane-bound protein, α-Syn has dramatically reduced affinity for, and does not tubulate, vesicles composed of a 1:1 POPG:POPC mixture. Coarse-grained molecular dynamics simulations suggested that the difference between the pure POPG and mixture results may be attributed to differences in the protein's partition depth, the membrane's hydrophobic thickness, and disruption of acyl chain order. To explore the importance of these attributes compared with the role of the reduced binding energy, we created an α-Syn 100 variant in which we removed the hydrophobic core of the non-amyloid component (NAC) domain and tested its impact on pure POPG vesicles. We observed a substantial reduction in binding affinity and tubulation, and simulations of the NAC-null protein suggested that the reduced binding energy increases the protein mobility on the bilayer surface, likely impacting the protein's ability to assemble into organized pretubule structures. We also used simulations to explore a potential role for interleaflet coupling as an additional driving force for tubulation. We conclude that symmetry across the leaflets in the tubulated state maximizes the interaction energy of the two leaflets and relieves the strain induced by the hydrophobic void beneath the amphipathic helix.
Membrane Shape Modulates Transmembrane Protein Distribution
Developmental Cell, 2014
Although membrane shape varies greatly throughout the cell, the contribution of membrane curvature to transmembrane protein targeting is unknown because of the numerous sorting mechanisms that take place concurrently in cells. To isolate the effect of membrane shape, we used cell-sized giant unilamellar vesicles (GUVs) containing either the potassium channel KvAP or the water channel AQP0 to form membrane nanotubes with controlled radii. Whereas the AQP0 concentrations in flat and curved membranes were indistinguishable, KvAP was enriched in the tubes, with greater enrichment in more highly curved membranes. Fluorescence recovery after photobleaching measurements showed that both proteins could freely diffuse through the neck between the tube and GUV, and the effect of each protein on membrane shape and stiffness was characterized using a thermodynamic sorting model. This study establishes the importance of membrane shape for targeting transmembrane proteins and provides a method for determining the effective shape and flexibility of membrane proteins.