Artificial Phospholipid Vesicles Research Articles

Analyzing the Interaction of Fluorescent-Labeled Proteins with Artificial Phospholipid Microvesicles using Quantitative Flow Cytometry.

In the human body, most of the major physiologic reactions involved in the immune response and blood coagulation proceed on the membranes of cells. An important first step in any membrane-dependent reaction is binding of protein on the phospholipid membrane. An approach to studying protein interaction with lipid membranes has been developed using fluorescently labeled proteins and flow cytometry. This method allows the study of protein-membrane interactions using live cells and natural or artificial phospholipid vesicles. The advantage of this method is the simplicity and availability of reagents and equipment. In this method, proteins are labeled using fluorescent dyes. However, both self-made and commercially available, fluorescently labeled proteins can be used. After conjugation with a fluorescent dye, the proteins are incubated with a source of the phospholipid membrane (microvesicles or cells), and the samples are analyzed by flow cytometry. The obtained data can be used to calculate the kinetic constants and equilibrium Kd. In addition, it is possible to estimate the approximate number of protein binding sites on the phospholipid membrane using special calibration beads.

Liposomes for the Delivery of Lipopeptide Vaccines.

Liposomes, which are artificial phospholipid vesicles with a bilayer membrane structure, have been developed and evaluated as a promising delivery system for vaccines. Here, we describe a procedure for the encapsulation of lipopeptide vaccines into liposomes. A liposomal formulation of lipid-core peptide was prepared via thin-film hydration followed by extrusion. The physicochemical properties of the liposomes, including their size, polydispersity, surface charge, and morphology, were analyzed using dynamic light scattering and transmission electron microscopy.

Pioglitazone Is a Mild Carrier-Dependent Uncoupler of Oxidative Phosphorylation and a Modulator of Mitochondrial Permeability Transition.

Pioglitazone (PIO) is an insulin-sensitizing antidiabetic drug, which normalizes glucose and lipid metabolism but may provoke heart and liver failure and chronic kidney diseases. Both therapeutic and adverse effects of PIO can be accomplished through mitochondrial targets. Here, we explored the capability of PIO to modulate the mitochondrial membrane potential (ΔΨm) and the permeability transition pore (mPTP) opening in different models in vitro. ΔΨm was measured using tetraphenylphosphonium and the fluorescent dye rhodamine 123. The coupling of oxidative phosphorylation was estimated polarographically. The transport of ions and solutes across membranes was registered by potentiometric and spectral techniques. We found that PIO decreased ΔΨm in isolated mitochondria and intact thymocytes and the efficiency of ADP phosphorylation, particularly after the addition of Ca2+. The presence of the cytosolic fraction mitigated mitochondrial depolarization but made it sustained. Carboxyatractyloside diminished the PIO-dependent depolarization. PIO activated proton transport in deenergized mitochondria but not in artificial phospholipid vesicles. PIO had no effect on K+ and Ca2+ inward transport but drastically decreased the mitochondrial Ca2+-retention capacity and protective effects of adenine nucleotides against mPTP opening. Thus, PIO is a mild, partly ATP/ADP-translocase-dependent, uncoupler and a modulator of ATP production and mPTP sensitivity to Ca2+ and adenine nucleotides. These properties contribute to both therapeutic and adverse effects of PIO.

Use of Thrombodynamics for revealing the participation of platelet, erythrocyte, endothelial, and monocyte microparticles in coagulation activation and propagation.

Background and objectiveFor many pathological states, microparticles are supposed to be one of the causes of hypercoagulation. Although there are some indirect data about microparticles participation in coagulation activation and propagation, the integral hemostasis test Thrombodynamics allows to measure micropaticles participation in these two coagulation phases directly. Demonstrates microparticles participation in coagulation activation by influence on the appearance of coagulation centres in the plasma volume and the rate of clot growth from the surface with immobilized tissue factor.Methods: Microparticles were obtained from platelets and erythrocytes by stimulation with thrombin receptor-activating peptide (SFLLRN) and calcium ionophore (A23187), respectively, from monocytes, endothelial HUVEC culture and monocytic THP cell culture by stimulation with lipopolysaccharides. Microparticles were counted by flow cytometry and titrated in microparticle-depleted normal plasma in the Thrombodynamics test.ResultsMonocyte microparticles induced the appearance of clotting centres through the TF pathway at concentrations approximately 100-fold lower than platelet and erythrocyte microparticles, which activated plasma by the contact pathway. For endothelial microparticles, both activation pathways were essential, and their activity was intermediate. Monocyte microparticles induced plasma clotting by the appearance of hundreds of clots with an extremely slow growth rate, while erythrocyte microparticles induced the appearance of a few clots with a growth rate similar to that from surface covered with high-density tissue factor. Patterns of clotting induced by platelet and endothelial microparticles were intermediate. Platelet, erythrocyte and endothelial microparticles impacts on the rate of clot growth from the surface with tissue factor did not differ significantly within the 0–200·103/ul range of microparticles concentrations. However, at concentrations greater than 500·103/ul, erythrocyte microparticles increased the stationary clot growth rate to significantly higher levels than do platelet microparticles or artificial phospholipid vesicles consisting of phosphatidylcholine and phosphatidylserine.ConclusionMicroparticles of different origins demonstrated qualitatively different characteristics related to coagulation activation and propagation.

The lipid droplet protein Pgc1 controls the subcellular distribution of phosphatidylglycerol.

The biosynthesis of yeast phosphatidylglycerol (PG) takes place in the inner mitochondrial membrane. Outside mitochondria, the abundance of PG is low. Here, we present evidence that the subcellular distribution of PG is maintained by the locally controlled enzymatic activity of the PG-specific phospholipase, Pgc1. A fluorescently labeled Pgc1 protein accumulates on the surface of lipid droplets (LD). We show, however, that LD are not only dispensable for Pgc1-mediated PG degradation, but do not even host any phospholipase activity of Pgc1. Our in vitro assays document the capability of LD-accumulated Pgc1 to degrade PG upon entry to the membranes of the endoplasmic reticulum, mitochondria and even of artificial phospholipid vesicles. Fluorescence recovery after photobleaching analysis confirms the continuous exchange of GFP-Pgc1 within the individual LD in situ, suggesting that a steady-state equilibrium exists between LD and membranes to regulate the immediate phospholipase activity of Pgc1. In this model, LD serve as a storage place and shelter Pgc1, preventing its untimely degradation, while both phospholipase activity and degradation of the enzyme occur in the membranes.

PH dependence of the charge recombination kinetics in bacterial RC reconstituted in liposomes

ABSTRACT:The photosynthetic Reaction Center from the carotenoidless mutant strain of the purple non sulphur bacterium Rhodobacter (R.) sphaeroides was reconstituted in artificial phospholipid vesicles (liposomes) to mimic the physiological membrane environment. The pH dependence in the interval 5 – 10 of the rate of the charge-recombination reactions from the final electron acceptors QA and QB to the primary electron donor (namely kAD and kBD) have been investigated. The liposomes were constituted of either the zwitterionic phosphatidylcholine (PC) or the negatively charged phosphatidylglycerol (PG), two of the main phospholipids found in the photosynthetic membrane of the bacterium. In both cases, the kAD has no pH dependence similarly to the detergent case. The kBD also has a pH dependence similar to the detergent case, having two distinct regions below pH 7 and above pH 9. Fitting of the titration curve to a function involving two protonation sites results in a marked shift of the pKAs between the different solubilizing environments. These differences are discussed in the frame of possible physiological implications.

Morphological Analysis of Trafficking and Processing of Anionic and Cationic Liposomes in Cultured Cells.

Liposomes, artificial phospholipid vesicles, have been developed as a non-viral drug delivery system to allow contained agents to be efficiently delivered to target sites via systemic circulation. Liposomes have been used as a gene transfer tool with cultured cells; however, their precise trafficking and processing remain uncertain. Furthermore, liposomes with different surface charges are known to exhibit distinct properties. The purpose of the current study was to elucidate the intracellular trafficking and processing of liposomes with anionic and cationic surface charges from a morphological view point. We found that cationic liposomes (CLs) were more effectively taken by the cells than anionic liposomes (ALs). Confocal laser scanning microscopy and transmission electron microscopy demonstrated distinct intracellular localization and processing patterns of ALs and CLs. ALs and their contents were localized in lysosomes but not in cytosol, indicating that ALs are subjected to the endosome-lysosome system. In contrast, contents of CLs were distributed mainly in the cytosol. CLs appear to disturb the cell membrane and then collapse to release their contents into the cytosol. It is feasible that the contents of CLs enter the cytosol directly rather than via the endosome-lysosome system.

In vitro spectroscopic study of piperine-encapsulated nanosize liposomes.

Black pepper is a source of effective antioxidants. It contains several powerful antioxidants and is thus one of the most important spices for preventing and curtailing oxidative stress. There is considerable interest in the development of a drug-delivery systems that would result in the selective delivery of antioxidants to tissues in sufficient concentrations to ameliorate oxidant-induced tissue injuries. Liposomes are biocompatible, biodegradable and nontoxic artificial phospholipid vesicles that offer the possibility of carrying hydrophilic, hydrophobic and amphiphilic molecules. This article focuses on the use of liposomes for the delivery of antioxidants in the prevention or treatment of pathological conditions related to oxidative stress. Liposome formulations of piperine were analyzed with various spectroscopic methods. The formulation with the highest entrapment efficiency (90.5%) was formulated with an L-α-phosphatidylcholine dipalmitoyl (DPPC):piperine, 30:1 molar ratio, and total lipid count of 19.47 mg/ml in the final liposomal preparation. The liposome formulation was found to be stable after storage at 4 °C, protected from light, for a minimum of 3 weeks. The incremental process of piperine penetration through the phospholipid membrane was analyzed using the FT-IR, UV-Vis and NMR methods. Temperature stability studies carried out at 37 °C showed the highest percentage of piperine release in the first 3 h of incubation.

Advances in missing-in-metastasis research

Missing-in-metastasis (MIM, also known as metastasis suppressor 1) belongs to the inverse BAR (I-BAR) family of intracellular membrane remodeling proteins. Structurally, MIM displays typical I-BAR family protein features including the ability to dimerize through its N-terminal I-BAR domain, which can be arrested by specific peptide inhibitors. It also induces filopodia-like membrane protrusions, and can reform artificial phospholipid vesicles. MIM binds to the inner plasma membrane and participates in multiple cellular activities including cell polarization, mobility, actin cytoskeleton regulation, protein interaction, cellular signal pathway facilitation, membrane shaping, and endocytosis. These activities are mostly performed in the nervous, circulatory, urinary, and reproductive systems. MIM abnormalities occur in a number of human diseases, especially those involving tumor genesis and metastasis. As well as MIM-specific peptides, RNA interference can be used to inhibit MIM expression. These molecular tools have the potential to be used in future bio-pharmaceutical developments. In this review, advances in MIM-related research are summarized, and the prospect of MIM targeting therapy and other MIM-based applications are introduced.

Liposomal antibiotic formulations for targeting the lungs in the treatment of Pseudomonas aeruginosa.

Pseudomonas aeruginosa is a Gram-negative bacterium that causes serious lung infections in cystic fibrosis, non-cystic fibrosis bronchiectasis, immunocompromised, and mechanically ventilated patients. The arsenal of conventional antipseudomonal antibiotic drugs include the extended-spectrum penicillins, cephalosporins, carbapenems, monobactams, polymyxins, fluoroquinolones, and aminoglycosides but their toxicity and/or increasing antibiotic resistance are of particular concern. Improvement of existing therapies against Pseudomonas aeruginosa infections involves the use of liposomes - artificial phospholipid vesicles that are biocompatible, biodegradable, and nontoxic and able to entrap and carry hydrophilic, hydrophobic, and amphiphilic molecules to the site of action. The goal of developing liposomal antibiotic formulations is to improve their therapeutic efficacy by reducing drug toxicity and/or by enhancing the delivery and retention of antibiotics at the site of infection. The focus of this review is to appraise the current progress of the development and application of liposomal antibiotic delivery systems for the treatment pulmonary infections caused by P.aeruginosa.

Nanoliposomes protect against AL amyloid light chain protein-induced endothelial injury

Context: A newly-recognized pathogenic mechanism underlying light chain amyloidosis (AL) involves endothelial dysfunction and cell injury caused by misfolded light chain proteins (LC). Nanoliposomes (NL) are artificial phospholipid vesicles that could attach to misfolded proteins and reduce tissue injury.Objective: To test whether co-treatment with NL reduces LC-induced endothelial dysfunction and cell death.Methods: Abdominal subcutaneous adipose arterioles from 14 non-AL subjects were cannulated; dilator response to acetylcholine and papaverine were measured at baseline and following 1-hour exposure to LC (20 µg/mL, 2 purified from AL subjects’ urine, 1 from human recombinant LC [AL-09]) ± NL (phosphatidylcholine/cholesterol/phosphatidic acid 70/25/5 molar ratio) or NL alone. Human aortic artery endothelial cells (HAEC) were exposed to Oregon Green-labeled LC ± NL for 24 hours and intracellular LC and apoptosis (Hoechst stain) were measured. Circular dichroism spectroscopy was performed on AL-09 LC ± NL to follow changes in secondary structure and protein thermal stability.Results: LC caused impaired dilation to acetylcholine that was restored by NL (control – 94.0 ± 1.8%, LC – 65.0 ± 7.1%, LC + NL – 95.3 ± 1.8%, p ≤ 0.001 LC versus control or LC + NL). NL protection was inhibited by L-NG-nitroarginine methyl ester. NL increased the beta sheet structure of LC, reduced endothelial cell internalization of LC and protected against LC-induced endothelial cell death.Conclusions: LC induced human adipose arteriole endothelial dysfunction and endothelial cell death, which were reversed by co-treatment with NL. This protection may partly be due to enhancing LC protein structure and reducing LC internalization. Nanoliposomes represent a promising new class of agents to ameliorate tissue injury from protein misfolding diseases such as AL.

Tau Oligomers Impair Artificial Membrane Integrity and Cellular Viability

The microtubule-associated protein Tau is mainly expressed in neurons, where it binds and stabilizes microtubules. In Alzheimer disease and other tauopathies, Tau protein has a reduced affinity toward microtubules. As a consequence, Tau protein detaches from microtubules and eventually aggregates into β-sheet-containing filaments. The fibrillization of monomeric Tau to filaments is a multistep process that involves the formation of various aggregates, including spherical and protofibrillar oligomers. Previous concepts, primarily developed for Aβ and α-synuclein, propose these oligomeric intermediates as the primary cytotoxic species mediating their deleterious effects through membrane permeabilization. In the present study, we thus analyzed whether this concept can also be applied to Tau protein. To this end, viability and membrane integrity were assessed on SH-SY5Y neuroblastoma cells and artificial phospholipid vesicles, treated with Tau monomers, Tau aggregation intermediates, or Tau fibrils. Our findings suggest that oligomeric Tau aggregation intermediates are the most toxic compounds of Tau fibrillogenesis, which effectively decrease cell viability and increase phospholipid vesicle leakage. Our data integrate Tau protein into the class of amyloidogenic proteins and enforce the hypothesis of a common toxicity-mediating mechanism for amyloidogenic proteins.

Liposomal Antioxidants for Protection against Oxidant-Induced Damage

Reactive oxygen species (ROS), including superoxide anion, hydrogen peroxide, and hydroxyl radical, can be formed as normal products of aerobic metabolism and can be produced at elevated rates under pathophysiological conditions. Overproduction and/or insufficient removal of ROS result in significant damage to cell structure and functions. In vitro studies showed that antioxidants, when applied directly and at relatively high concentrations to cellular systems, are effective in conferring protection against the damaging actions of ROS, but results from animal and human studies showed that several antioxidants provide only modest benefit and even possible harm. Antioxidants have yet to be rendered into reliable and safe therapies because of their poor solubility, inability to cross membrane barriers, extensive first-pass metabolism, and rapid clearance from cells. There is considerable interest towards the development of drug-delivery systems that would result in the selective delivery of antioxidants to tissues in sufficient concentrations to ameliorate oxidant-induced tissue injuries. Liposomes are biocompatible, biodegradable, and nontoxic artificial phospholipid vesicles that offer the possibility of carrying hydrophilic, hydrophobic, and amphiphilic molecules. This paper focus on the use of liposomes for the delivery of antioxidants in the prevention or treatment of pathological conditions related to oxidative stress.

Control of Respiration by Cytochrome c Oxidase in Intact Cells

Metabolic control analysis was applied to intact HepG2 cells. The effect on the control coefficient of cytochrome c oxidase (CcOX) over cell respiration of both the electrical (Delta psi) and chemical (Delta pH) component of the mitochondrial transmembrane proton electrochemical gradient (Delta mu(H(+))) was investigated. The overall O(2) consumption and specific CcOX activity of actively phosphorylating cells were titrated with cyanide under conditions in which Delta psi and Delta pH were selectively modulated by addition of ionophores. In the absence of ionophores, CcOX displayed a high control coefficient (C(IV) = 0.73), thus representing an important site of regulation of mitochondrial oxidative phosphorylation. A high control coefficient value (C(IV) = 0.85) was also measured in the presence of nigericin, i.e. when Delta psi is maximal, and in the presence of nigericin and valinomycin (C(IV) = 0.77), when Delta mu(H(+)) is abolished. In contrast, CcOX displayed a markedly lower control coefficient (C(IV) = 0.30) upon addition of valinomycin, when Delta psi is converted into Delta pH. These results show that Delta psi is responsible for the tight control of CcOX over respiration in actively phosphorylating cells.

Interorganelle Trafficking of Ceramide Is Regulated by Phosphorylation-dependent Cooperativity between the PH and START Domains of CERT

The synthesis and transport of lipids are essential events for membrane biogenesis. However, little is known about how intracellular trafficking of lipids is regulated. Ceramide is synthesized at the endoplasmic reticulum (ER) and transported by the ceramide transfer protein CERT to the Golgi apparatus, where it is converted to sphingomyelin. CERT has a phosphoinositide-binding pleckstrin homology (PH) domain for Golgi-targeting and a lipid transfer START domain for intermembrane transfer of ceramide. We here show that CERT receives multiple phosphorylations at a serine-repeat motif, a possibe site for casein kinase I, and that the phosphorylation down-regulates the ER-to-Golgi transport of ceramide. In vitro assays show that the phosphorylation induces an autoinhibitory interaction between the PH and START domains and consequently inactivates both the phosphoinositide binding and ceramide transfer activities of CERT. Loss of sphingomyelin and cholesterol from cells causes dephosphorylation of CERT to activate it. The cooperative control of functionally distinct domains of CERT is a novel molecular event to regulate the intracellular trafficking of ceramide.

A Novel Mechanism of Interaction between α-Synuclein and Biological Membranes

Conformational abnormalities and aggregation of α-synuclein (α-syn) have been linked to the pathogenesis of Parkinson’s (PD) and related diseases. It has been shown that α-syn can stably bind artificial phospholipid vesicles through α-helix formation in its N-terminal repeat region. However, little is known about the membrane interaction in cells. In the current study, we determined the membrane-binding properties of α-syn to biological membranes by using bi-functional chemical crosslinkers, which allow the detection of transient, but specific, interactions. By utilizing various point mutations and deletions within α-syn, we demonstrated that the membrane interaction of α-syn in cells is also mediated by α-helix formation in the N-terminal repeat region. Moreover, the PD-linked A30P mutation causes reduced membrane binding, which is concordant with the artificial membrane studies. However, contrary to the interaction with artificial membranes, the interaction with biological membranes is rapidly reversible and is not driven by electrostatic attraction. Furthermore, the interaction of α-syn with cellular membranes occurs only in the presence of non-protein and non-lipid cytosolic components, which distinguishes it from the spontaneity of the interaction with artificial membranes. More interestingly, addition of the cytosolic preparation to artificial membranes resulted in the transient, charge-independent binding of α-syn similar to the interaction with biological membranes. These results suggest that in cells, α-syn is engaged in a fundamentally different mode of membrane interaction than the charge-dependent artificial membrane binding, and the mode of interaction is determined by the intrinsic properties of α-syn itself and by the cytoplasmic context.

Molten globule structure and steroidogenic activity of N-218 MLN64 in human placental mitochondria.

Progesterone synthesis by the human placenta requires the conversion of mitochondrial cholesterol to pregnenolone by cytochrome P450scc. Most steroidogenic tissues use the steroidogenic acute regulatory protein (StAR) to deliver cholesterol to the inner mitochondrial membrane where P450scc is located, but StAR is not expressed in the human placenta. However, the human placenta does express MLN64, which has a C-terminal domain homologous to StAR that can also transport cholesterol. We investigated the ability of bacterially expressed N-218 MLN64 and N-62 StAR to transport cholesterol between artificial membranes and to its inner membrane site of use in placental mitochondria. Urea denaturation experiments show that N-218 MLN64 undergoes a pH-dependent and denaturant-dependent structural transition to a molten globule state, as reported previously for N-62 StAR. N-218 MLN64 stimulated cholesterol transfer between artificial phospholipid vesicles with an initial rate of 6.5 mol/min.mol N-218 MLN64. Both N-218 MLN64 and N-62 StAR stimulated cholesterol transfer to the inner mitochondrial membrane, as evidenced by a 6-fold stimulation of pregnenolone synthesis with saturating transporter. This stimulation was seen only after the endogenous cholesterol in the steroidogenic pool of the isolated mitochondria was first depleted. No stimulation was observed by N-218 MLN64 or N-62 StAR when 20alpha-hydroxycholesterol was added as substrate for P450scc, confirming that these proteins stimulate P450scc activity by enhancing cholesterol transport. MLN64 levels in placental JEG-3 cells were unresponsive to stimulation by 8-bromo-cAMP over 24 h. These data show that human N-218 MLN64 and N-62 StAR have similar biophysical and functional properties and are able to stimulate steroidogenesis in a human placental system, which normally lacks StAR. The results reveal that with saturating MLN64, steroidogenesis by placental mitochondria proceeds at near-maximal rate.

Assay for G protein-dependent activation of phospholipase C beta using purified protein components.

The activity of mammalian phosphoinositide-specific phospholipase C beta (PLC beta) is regulated by the alpha q family of G protein alpha subunits and by beta gamma subunits thought to be released from Gi. Interactions between G protein subunits and PLC beta can be assayed by measuring the stimulation of PLC beta enzymatic activity on reconstituting the purified G protein subunits with purified PLC beta on artificial phospholipid vesicles containing the substrate, phosphatidylinositol-4,5-bisphosphate (PIP2). These vesicles are doped with [3H]-inositol PIP2 and the rate of hydrolysis is determined by quantitating the amount of [3H]-inositol triphosphate (IP3) released from the vesicle into the aqueous phase. This assay provides a relatively simple method for assessing the activity PLC activity and its ability to be regulated by beta gamma and alpha(q) subunits. It can also be used to assess the functionality of the components after modification by mutagenesis, chemical modification, or in the presence of competing molecules.

Activation of Phosphoinositide 3-Kinase γ by Ras

Background: Type I phosphoinositide 3-kinases are responsible for the hormone-sensitive synthesis of the lipid messenger phosphatidylinositol(3,4,5)-trisphosphate. Type IA and IB subfamily members contain a Ras binding domain and are stimulated by activated Ras proteins both in vivo and in vitro. The mechanism of Ras activation of type I PI3Ks is unknown, in part because no robust in vitro assay of this event has been established and characterized. Other Ras effectors, such as Raf and phosphoinositide-phospholipase Cϵ, have been shown to be translocated into the plasma membrane, leading to their activation.Results: We show that posttranslationally lipid-modified, activated N-, H-, K-, and R-Ras proteins can potently and substantially activate PI3Kγ when using a stripped neutrophil membrane fraction as a source of phospholipid substrate. We have found GTPγS-loaded Ras can significantly (6- to 8-fold) activate PI3Kγ when using artificial phospholipid vesicles containing their substrate, and this effect is a result of both a decrease in apparent Km for phosphatidylinositol(4,5)-bisphosphate and an increase in the apparent Vmax. However, neither in vivo nor in the two in vitro assays of Ras activation of PI3Kγ could we detect any evidence of a Ras-dependent translocation of PI3Kγ to its source of phospholipid substrate.Conclusions: Our data suggest that Ras activate PI3Kγ at the level of the membrane, by allosteric modulation and/or reorientation of the PI3Kγ, implying that Ras can activate PI3Kγ without its membrane translocation. This view is supported by structural work that has suggested binding of Ras to PI3Kγ results in a change in the structure of the catalytic pocket.

How helminth lipid-binding proteins offload their ligands to membranes: differential mechanisms of fatty acid transfer by the ABA-1 polyprotein allergen and Ov-FAR-1 proteins of nematodes and Sj-FABPc of schistosomes.

Three different classes of small lipid-binding protein (LBP) are found in helminth parasites. Although of similar size, the ABA-1A1 (also designated As-NPA-A1) and Ov-FAR-1 (formerly known as Ov20) proteins of nematodes are mainly alpha-helical and have no known structural counterparts in mammals, whereas Sj-FABPc of schistosomes is predicted to form a beta-barrel structure similar to the mammalian family of intracellular fatty acid binding proteins. The parasites that produce these proteins are unable to synthesize their own complex lipids and, instead, rely entirely upon their hosts for supply. As a first step in elucidating whether these helminth proteins are involved in the acquisition of host lipid, the process by which these LBPs deliver their ligands to acceptor membranes was examined, by comparing the rates and mechanisms of ligand transfer from the proteins to artificial phospholipid vesicles using a fluorescence resonance energy transfer assay. All three proteins bound the fluorescent fatty acid 2-(9-anthroyloxy)palmitic acid (2AP) similarly, but there were clear differences in the rates and mechanisms of fatty acid transfer. Sj-FABPc displayed a collisional mechanism; 2AP transfer rates increased with acceptor membrane concentration, were modulated by acceptor membrane charge, and were not diminished in the presence of increasing salt concentrations. In contrast, transfer of ligand from Ov-FAR-1 and ABA-1A1 involved an aqueous diffusion step; transfer rates from these proteins were not modulated by acceptor membrane concentration or charge but did decrease with the ionic strength of the buffer. Despite these differences, all of the proteins interacted directly with membranes, as determined using a cytochrome c competition assay, although Sj-FABPc interacted to a greater extent than did Ov-FAR-1 or rABA-1A1. Together, these results suggest that Sj-FABPc is most likely to be involved in the intracellular targeted transport and metabolism of fatty acids, whereas Ov-FAR-1 and ABA-1A1 may behave in a manner analogous to that of extracellular LBPs such as serum albumin and plasma retinol binding protein.