Phase diagram of a polyunsaturated lipid mixture: Brain sphingomyelin/1-stearoyl-2-docosahexaenoyl-sn-glycero-3-phosphocholine/cholesterol

Konyakhina, Tatyana M.; Feigenson, Gerald W.

doi:10.1016/j.bbamem.2015.10.016

Cited by 52 publications

(59 citation statements)

References 105 publications

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“…Along a tieline the fraction of each coexisting phase is known. Currently, the majority of published phase diagrams describe neutral lipid mixtures (36)(37)(38)(39)48). No published data are available with ternary phase diagrams containing PC/PS/Chol.…”

Section: Resultsmentioning

confidence: 99%

“…One previous study found a nonphysiological enrichment of cholesterol in virions compared to the cellular plasma membrane (33); however, a more thorough study found no significant enrichment in cholesterol (35). Additionally, in model membranes with coexisting raft and non-raft domains at a physiological cholesterol mole fraction of 0.35 to 0.4, cholesterol is present at similar levels in the coexisting domains (36)(37)(38)(39). Also, under some conditions Gag is found in cold detergent-resistant membranes (DRMs) along with raft markers (40)(41)(42)(43).…”

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confidence: 92%

“…Light scattering and fluorescence background were controlled as described previously (71, 72). Lipids were mixed in chloroform to 120 nmol, and the organic solvent was replaced by buffer using rapid solvent exchange (RSE) (68) and then sealed GUV samples described here were prepared by gentle hydration (38,73). A total of 300 nmol total lipid containing 0.02 mol% LR-DOPE was mixed in chloroform, partially dried to a thin film in a culture tube using a rotary evaporator, and then dried with heating at 55°C under high vacuum for 1.5 h. The thin dry film was then hydrated with wet N 2 gas at 55°C for 30 min.…”

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confidence: 99%

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Effects of Membrane Charge and Order on Membrane Binding of the Retroviral Structural Protein Gag

Wen

Dick

Feigenson

et al. 2016

J Virol

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The retroviral structural protein Gag binds to the inner leaflet of the plasma membrane (PM), and many cellular proteins do so as well. We used Rous sarcoma virus (RSV) Gag together with membrane sensors to study the principles governing peripheral protein membrane binding, including electrostatics, specific recognition of phospholipid headgroups, sensitivity to phospholipid acyl chain compositions, preference for membrane order, and protein multimerization. We used an in vitro liposome-pelleting assay to test protein membrane binding properties of Gag, the well-characterized MARCKS peptide, a series of fluorescent electrostatic sensor proteins (mNG-KRn), and the specific phosphatidylserine (PS) binding protein Evectin2. RSV Gag and mNG-KRn bound well to membranes with saturated and unsaturated acyl chains, whereas the MARCKS peptide and Evectin2 preferentially bound to membranes with unsaturated acyl chains. To further discriminate whether the primary driving force for Gag membrane binding is electrostatic interactions or preference for membrane order, we measured protein binding to giant unilamellar vesicles (GUVs) containing the same PS concentration in both disordered (Ld) and ordered (Lo) phases. RSV Gag and mNG-KRn membrane association followed membrane charge, independent of membrane order. Consistent with pelleting data, the MARCKS peptide showed preference for the Ld domain. Surprisingly, the PS sensor Evectin2 bound to the PS-rich Ld domain with 10-fold greater affinity than to the PS-rich Lo domain. In summary, we found that RSV Gag shows no preference for membrane order, while proteins with reported membrane-penetrating domains show preference for disordered membranes. IMPORTANCERetroviral particles assemble on the PM and bud from infected cells. Our understanding of how Gag interacts with the PM and how different membrane properties contribute to overall Gag assembly is incomplete. This study examined how membrane charge and membrane order influence Gag membrane association. Consistent with previous work on RSV Gag, we report here that electrostatic interactions provide the primary driving force for RSV Gag membrane association. Using phase-separated GUVs with known lipid composition of the Ld and Lo phases, we demonstrate for the first time that RSV Gag is sensitive to membrane charge but not membrane order. In contrast, the cellular protein domain MARCKS and the PS sensor Evectin2 show preference for disordered membranes. We also demonstrate how to define GUV phase composition, which could serve as a tool in future studies of protein membrane interactions.T he retroviral structural protein Gag provides the primary driving force for virus assembly at the inner leaflet of the plasma membrane (PM), and Gag alone is sufficient to assemble into virus-like particles (VLPs) in cells (1) and also in vitro (2). Gag is synthesized in the cytosol and then is targeted to the PM by the N-terminal domain, MA. How Gag interacts with the inner leaflet of the PM and how the biophysical properties of the PM...

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Section: Resultsmentioning

confidence: 99%

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confidence: 92%

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confidence: 99%

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Effects of Membrane Charge and Order on Membrane Binding of the Retroviral Structural Protein Gag

Wen

Dick

Feigenson

et al. 2016

J Virol

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“…To do this, we prepared model membrane vesicles composed of various combinations of B. burgdorferi lipids. Model membrane vesicles have proven to be very useful systems for defining the abilities of lipids to form ordered domains in membranes and identifying conditions in which such domains exist as nanoscale-size domains (38)(39)(40)(41)(42)(43)(44)(45). Using model membrane vesicles, we found that the major Borrelia lipids differ greatly in their ability to support ordered-domain/raft formation.…”

Section: Introductionmentioning

confidence: 90%

Ordered Membrane Domain-Forming Properties of the Lipids of Borrelia burgdorferi

Huang

Toledo

Benach

et al. 2016

Biophysical Journal

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Co-existing disordered and ordered (raft) membrane domains exist in Borrelia burgdorferi, the causative agent of Lyme disease. However, although B. burgdorferi contains cholesterol lipids, it lacks sphingolipids-a crucial component of rafts in eukaryotes. To define the principles of ordered lipid domain formation in Borrelia, the domain forming properties of vesicles composed of its three major lipids, acylated cholesteryl galactoside (ACGal), monogalactosyl diacyglycerol (MGalD), and phosphatidylcholine (PC) and/or their mixtures were studied. Anisotropy and fluorescence resonance energy transfer measurements were used to assay membrane order and ordered-domain formation. ACGal had the highest potential to form ordered domains. Interestingly, mixtures of ACGal with B. burgdorferi PC formed ordered domains more readily than mixtures of ACGal with MGalD. This appears to reflect the relatively high level of saturation observed for B. burgdorferi PC, as vesicles containing ACGal and PC, but in which the unsaturated lipid dioleoyl PC was substituted for Borrelia PC, failed to form ordered domains. In addition, the properties of ACGal were compared to those of cholesterol. Depending on what other lipids were present, ordered-domain formation in the presence of ACGal was greater than or equal to that in the presence of cholesterol. Giant unilamellar vesicles formed from ACGal-containing mixtures showed rounded domain shapes similar to those in analogous vesicles containing cholesterol, indicative of liquid-ordered state rather than solid-like gel-state domain formation. Over all, principles of ordered-domain formation in B. burgdorferi appear to be very similar to those in eukaryotes, with saturated PC taking the place of sphingolipids, but with ACGal being the main lipid component inducing ordered-domain formation.

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“…In particular, >60% of the non-cholesterol lipids are asymmetric, meaning they contain one polyunsaturated fatty acid (PUFA) and one saturated fatty acid; cholesterol makes up 45% of the total lipid content; and sphingomyelin makes up just 4% of the composition [10,11]. Konyakhina & Feigenson recently solved a phase diagram for a mixture containing asymmetric PUFA-containing lipids, sphingomyelin, and cholesterol [12]. In the region of Lo/Ld phase coexistence, these PUFA-containing lipids form the Ld phase while sphingomyelin and cholesterol pack into the Lo phase, much like a canonical raft mixture.…”

Section: Introductionmentioning

confidence: 99%

Polyunsaturated chains in asymmetric lipids disorder raft mixtures and preferentially associate with α-Synuclein

Brummel¹,

Braun²,

Sachs³

2017

Biochimica et Biophysica Acta (BBA) - Biomembranes

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Using molecular dynamics simulations, we have explored the effect of asymmetric lipids—specifically those that contain one polyunsaturated (PUFA) and one saturated fatty acid chain—on phase separation in heterogeneous membranes. These lipids are prevalent in neuronal membranes, particularly in synaptic membranes, where the Parkinson's Disease protein α-Synuclein (αS) is found. We have therefore explored the relationship between asymmetric, PUFA-containing lipids, and αS. The simulations show that asymmetric lipids partition to the liquid disordered (Ld) phase of canonical raft mixtures because of the highly disordered PUFA chain. In the case of a membrane built to mimic the lipid composition of a synaptic vesicle, the PUFA-containing asymmetric lipids completely disrupt phase separation. Because αS is positively charged, we show that it partitions with negatively charged lipids, regardless of the saturation state of the chains. Additionally, αS preferentially associates with the polyunsaturated fatty acid tails of both charged and neutral lipids. This is a consequence of those chains' ability to accommodate the void beneath the amphipathic helix. This article is part of a Special Issue entitled: Lipid order/lipid defects and lipid-control of protein activity edited by Dirk Schneider.

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Phase diagram of a polyunsaturated lipid mixture: Brain sphingomyelin/1-stearoyl-2-docosahexaenoyl-sn-glycero-3-phosphocholine/cholesterol

Cited by 52 publications

References 105 publications

Effects of Membrane Charge and Order on Membrane Binding of the Retroviral Structural Protein Gag

Effects of Membrane Charge and Order on Membrane Binding of the Retroviral Structural Protein Gag

Ordered Membrane Domain-Forming Properties of the Lipids of Borrelia burgdorferi

Polyunsaturated chains in asymmetric lipids disorder raft mixtures and preferentially associate with α-Synuclein

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