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At least 217 records · Page 12

Dietary n-3 polyunsaturated fatty acids affect the development of renovascular hypertension in rats

The consequences of a dietary n-3 PUFA supply was investigated on the blood pressure (BP) increase elicited by left renal artery stenosis in rats distributed in 3 groups (n = 8) fed for 8 weeks a semi-purified diet either as control diet or enriched diets (docosahexaenoic acid, DHA, or eicosapentaenoic acid, EPA). The PUFA intake induced large alterations in heart and kidney phospholipid fatty acid profile, but did not influence body weight, cardiac hypertrophy, renal left atrophy and right hypertrophy. Within 4 weeks, BP raised from 120-180 +/- 2 mm Hg in the control group, but only to 165 +/- 3 mm Hg in the n-3 PUFA groups. After stabilization of BP in the 3 groups, the rats received a short administration of increasing dose of perindopril. The lower dose (0.5 mg/kg) moderately decreased BP only in the control group. With higher doses (1, 5 and 10 mg/kg) BP was normalized in the 3 groups, with a higher amplitude of the BP lowering effect in the control group. A moderate n-3 PUFA intake can contribute to prevent the development of peripheral hypertension in rats by a mechanism that may involve angiotensin converting enzyme.

Hypertension, Renovascular/etiology/pathology/phys↗

Quantitative determination of microbial activity and community nutritional status in estuarine sediments: evidence for a disturbance artifact

In estuarine sediments with a high degree of vertical heterogeneity in reduced substrate and terminal electron acceptor concentrations, the method of exposure of the microbiota to labeled substrates can introduce a "disturbance artifact" into measures of metabolic activity. The detection of this artifact is based on quantitative measurement of the relative rates of incorporation of [14C]acetate into phospholipid fatty acids (PLFA) and endogenous storage lipid, poly-beta-hydroxyalkanoate (PHA). Previous studies have shown that PLFA synthesis measures cellular growth and that PHA synthesis measures carbon accumulation (unbalanced growth). The "disturbance artifact" of exposure to [14C]acetate was demonstrated by comparing injection of a core with the usual or pore-water replacement or slurry techniques. Only injection of labeled substrate allowed detection of preassay disturbance of the sediment with a garden rake. The raking increased PLFA synthesis with little effect to differences in concentration or distribution of [14C]acetate in the 10-min incubation. Bioturbation induced by sand dollar feeding in estuarine sediment could be detected in an increased PLFA/PHA ratio which was due to decreased PHA synthesis if the addition of labeled substrate was by the injection technique. Addition of labeled precursors to sediment by slurry or pore-water replacement induces greater disturbance artifacts than injection techniques.

Phospholipids/analysis/biosynthesis↗

Computer simulation of ion channel gating: the M(2) channel of influenza A virus in a lipid bilayer

The transmembrane fragment of the influenza virus M(2) protein forms a homotetrameric channel that transports protons. In this paper, we use molecular dynamics simulations to help elucidate the mechanism of channel gating by four histidines that occlude the channel lumen in the closed state. We test two competing hypotheses. In the "shuttle" mechanism, the delta nitrogen atom on the extracellular side of one histidine is protonated by the incoming proton, and, subsequently, the proton on the epsilon nitrogen atom is released on the opposite side. In the "water-wire" mechanism, the gate opens because of electrostatic repulsion between four simultaneously biprotonated histidines. This allows for proton transport along the water wire that penetrates the gate. For each system, composed of the channel embedded in a hydrated phospholipid bilayer, a 1.3-ns trajectory was obtained. It is found that the states involved in the shuttle mechanism, which contain either single-protonated histidines or a mixture of single-protonated histidines plus one biprotonated residue, are stable during the simulations. Furthermore, the orientations and dynamics of water molecules near the gate are conducive to proton transfer. In contrast, the fully biprotonated state is not stable. Additional simulations show that if only two histidines are biprotonated, the channel deforms but the gate remains closed. These results support the shuttle mechanism but not the gate-opening mechanism of proton gating in M(2).

NASA Center ARC↗

Phosphoinositide kinases and the synthesis of polyphosphoinositides in higher plant cells

Phosphoinositides are a family of inositol-containing phospholipids which are present in all eukaryotic cells. Although in most cells these lipids, with the exception of phosphatidylinositol, constitute only a very minor proportion of total cellular lipids, they have received immense attention by researchers in the past 15-20 years. This is due to the discovery that these lipids, rather than just having structural functions, play key roles in a wide range of important cellular processes. Much less is known about the plant phosphoinositides than about their mammalian counterparts. However, it has been established that a functional phosphoinositide system exists in plant cells and it is becoming increasingly clear that inositol-containing lipids are likely to play many important roles throughout the life of a plant. It is not our intention to give an exhaustive overview of all aspects of the field, but rather we focus on the phosphoinositide kinases responsible for the synthesis of all phosphorylated forms of phosphatidylinositol. Also, we mention some of the aspects of current phosphoinositide research which, in our opinion, are most likely to provide a suitable starting point for further research into the role of phosphoinositides in plants.

Review↗

Transient and sustained increases in inositol 1,4,5-trisphosphate precede the differential growth response in gravistimulated maize pulvini

The internodal maize pulvinus responds to gravistimulation with differential cell elongation on the lower side. As the site of both graviperception and response, the pulvinus is an ideal system to study how organisms sense changes in orientation. We observed a transient 5-fold increase in inositol 1,4,5-trisphosphate (IP3) within 10 s of gravistimulation in the lower half of the pulvinus, indicating that the positional change was sensed immediately. Over the first 30 min, rapid IP3 fluctuations were observed between the upper and lower halves. Maize plants require a presentation time of between 2 and 4 h before the cells on the lower side of the pulvinus are committed to elongation. After 2 h of gravistimulation, the lower half consistently had higher IP3, and IP3 levels on the lower side continued to increase up to approximately 5-fold over basal levels before visible growth. As bending became visible after 8-10 h, IP3 levels returned to basal values. Additionally, phosphatidylinositol 4-phosphate 5-kinase activity in the lower pulvinus half increased transiently within 10 min of gravistimulation, suggesting that the increased IP3 production was accompanied by an up-regulation of phosphatidylinositol 4, 5-bisphosphate biosynthesis. Neither IP3 levels nor phosphatidylinositol 4-phosphate 5-kinase activity changed in pulvini halves from vertical control plants. Our data indicate the involvement of IP3 and inositol phospholipids in both short- and long-term responses to gravistimulation. As a diffusible second messenger, IP3 provides a mechanism to transmit and amplify the signal from the perceiving to the responding cells in the pulvinus, coordinating a synchronized growth response.

NASA Discipline Plant Biology↗

N-methyl-D-aspartate increases acetylcholine release from rat striatum and cortex: its effect is augmented by choline

We examined the effects of N-methyl-D-aspartate (NMDA), a glutamate agonist, and of glutamate itself, on acetylcholine (ACh) release from superfused rat striatal slices. In a Mg(++)-free medium, NMDA (32-1000 microM) as well as glutamate (1 mM) increased basal ACh release by 35 to 100% (all indicated differences, P less than .05), without altering tissue ACh or choline contents. This augmentation was blocked by Mg++ (1.2 mM) or by MK-801 (10 microM). Electrical stimulation (15 Hz, 75 mA) increased ACh release 9-fold (from 400 to 3660 pmol/mg of protein): this was enhanced (to 4850 pmol/mg of protein) by NMDA (100 microM). ACh levels in stimulated slices fell by 50 or 65% depending on the absence or presence of NMDA. The addition of choline (40 microM) increased ACh release both basally (570 pmol/mg of protein) and with electrical stimulation (6900 pmol/mg of protein). In stimulated slices choline acted synergistically with NMDA, raising ACh release to 10,520 pmol/mg of protein. The presence of choline also blocked the fall in tissue ACh. No treatment affected tissue phospholipid or protein levels. NMDA (32-320 microM) also augmented basal ACh release from cortical but not hippocampal slices. Choline efflux from striatal and cortical (but not hippocampal) slices decreased by 34 to 50% in Mg(++)-free medium. These data indicate that NMDA-like drugs may be useful, particularly in combination with choline, to enhance striatal and cortical cholinergic activity. ACh release from rat hippocampus apparently is not affected by NMDA receptors.

Non-NASA Center↗

Bacterial Composition and Quantitation in the Yungay Region of the Atacama Desert, Chile

Voyager would have produced similar negative results if tested in the Yungay region (Navarro-Gonzales et al, 2003). However, the number of bacteria present per gr'am in the Yungay sands is not well-established, being below the detection limit of culture methods. For the past several years, this group has been analyzing Yungay sands by quantitative Polymerase Chain Reaction (qPCR), both in the lab and in the field, to establish protocols for counting bacteria. We use primers that detect a broad range of bacterial taxa, including both proteobacteria and firmicutes. Optimization of DNA extraction and qPCR analysis is critical for accurate counts. Comparison of qPCR results with PhosphoLipid Fatty Acid (PLFA)-analysis and other counting methods indicates that there are detectable bacteria in sand obtained from the upper few centimeters of the desert surface.

Fletcher, Lauren↗

A Radical Pathway for Organic Phosphorylation during Schreibersite Corrosion with Implications for the Origin of Life

Phosphorylated compounds (e.g. DNA, RNA, phospholipids, and many coenzymes) are critical to biochemistry. Thus, their origin is of prime interest to origin of life studies. The corrosion of the meteoritic mineral schreibersite ((Fe,Ni)3P) may have significantly contributed to the origin of phosphorylated biomolecules. Corrosion of synthetic schreibersite in a variety of solutions was analyzed by nuclear magnetic resonance spectroscopy, mass spectrometry, and electron paramagnetic resonance spectroscopy. These methods suggest a radical reaction pathway for the corrosion of schreibersite to form phosphite radicals (raised dot PO3 sup 2-)) aqueous solution. These radicals can form activated polyphosphates and can phosphorylate organic compounds such as acetate (3% yield). Phosphonates (O3P-C) are found in the organic P inventory of the carbonaceous meteorite Murchison. While phosphonates are rare in biochemistry, the ubiquity of corroding iron meteorites on the early Earth could have provided an accessible source of organophosphorous for the origin of life allowing the invention of the organophosphates in modern biology as a product of early evolution.

Pasek, Matthew A.↗

Processes that Drove the Transition from Chemistry to Biology: Concepts and Evidence

Two properties are particularly germane to the transition from chemistry to biology. One is the emergence of complex molecules (polymers) capable of performing non-trivial functions, such as catalysis, energy transduction or transport across cell walls. The other is the ability of several functions to work in concert to provide reproductive advantage to systems hosting these functions. Biological systems exhibit these properties at remarkable levels of efficiency and accuracy in a way that appears effortless. However, dissection of these properties reveals great complexities that are involved. This opens a question: how a simple, ancestral system could have acquired the required properties? Other questions follow. What are the chances that a functional polymer emerges at random? What is the minimum structural complexity of a polymer to carry out a function at a reasonable level of efficiency? Can we identify concrete, protobiologically plausible mechanisms that yield advantageous coupling between different functions? These and similar questions are at the core of the main topic of this session: how soulless chemistry became life? Clearly, we do not have complete answers to any of these questions. However, in recent years a number of new and sometimes unexpected clues have been brought to light. Of particular interest are proteins because they are the main functional polymers in contemporary cells. The emergence of protein functions is a puzzle. It is widely accepted that a well ]defined, compact structure (fold) is a prerequisite for function. It is equally widely accepted that compact folds are rare among random amino acid polymers. Then, how did protein functionality start? According to one hypothesis well folded were preceded by their poorly folded, yet still functional ancestors. Only recently, however, experimental evidence supporting this hypothesis has been presented. In particular, a small enzyme capable of ligating two RNA fragments with the rate of 106 above background was evolved in vitro. This enzyme does not look like any contemporary protein. It is very flexible and its structure is kept together just by a single salt bridge between a charged residue and a coordinating zinc. A similar picture emerges from studies of simple transmembrane channels that mimic those in ancestral cells. Again, they are extremely flexible and do not form a conventional pore. Yet, they efficiently mediate ion transport. Studies on simple proteins that are on-going in several laboratories hold promise of revealing crucial links between chemical and biological catalysis and other ubiquitous cell functions. Interaction between composition, growth and division of protobiologically relevant vesicles and metabolic reactions that they encapsulate is an example of coupling between simple functions that promotes reproduction and evolution. Recent studies have demonstrated possible mechanisms by which vesicles might have evolved their composition from fatty acids to phospholipids, thus facilitating a number of new cellular functions. Conversely, it has been also demonstrated that an encapsulated metabolism might drive vesicle division. These are, again, examples of processes that might have driven the transition from chemistry to biology.

Pohorille, Andrew↗

Microbial Lipid and C Isotopic Biosignatures of a Unique Community at Grand Prismatic Spring, Yellowstone National Park

The microbial communities found in modern hot springs are considered analogs to ones that may have existed in hydrothermal systems on the early Earth and possibly Mars. Our goal was to characterize the microbial biosignatures and to assess the preservation of organic matter in the silica-depositing Grand Prismatic Spring in Yellowstone National Park. This study combines 16S rRNA surveys, lipid biomarkers, and C isotopes to query, "Who's there and what are they doing?" On the edge of the approximately 90 m diameter blue vent pool (56.1 C, pH 8.5), a floating green streamer community grew over a benthic pink community. The membrane lipids in the green streamers and pink mat were composed of unusual ester-linked fatty acids, indicating the presence of novel bacterial groups. In particular, we discovered a series of 2-methyl and 2,X-dimethyl phospholipid fatty acids (C18-22). We are attempting to use the 16S rRNA surveys to link these compounds to source organisms. Wax esters, biomarkers for Chloroflexi, were present in both communities, but displayed different profiles. A higher proportion of branched wax esters were found in the green streamers, and were associated with a relatively high concentration of long-chain di- and trienes (C29-31). This suggests that Chloroflexus primarily grew in the green streamers, while a pink mat of Roseiflexus grew on the sinter substrate underneath. Cyanobacterial alkanes were found in the green streamers (n-C17, 7-, 6- and 5-monomethyl-C17, 7,11-dimethyl-C17, n-C19, n-C19:1). We also detected a series of monoalkylglycerylethers and geologically relevant hopanoids in both communities. Carbon isotope analyses indicated that Chloroflexus was growing photoheterotrophically using cyanobacterial photosynthate. Roseiflexus also traditionally grows photoheterotrophically, but the C isotopic signatures of the lipids in the pink mat were approximately 10 %0 lighter than the cyanobacterial and Chloroflexus lipids, indicating a potentially novel metabolic mechanism or prior secondary reworking of substrates before reaching Roseiflexus. This arrangement of Synechococcus-Chloroflexus green streamers floating over a benthic pink community of Roseiflexus is different from the classical laminated Synechococcus-Chloroflexi mats at Octopus Hot Spring.

Jahnke, Linda↗

Microorganisms, Organic Carbon, and Their Relationship with Oxidant Activity in Hyper-Arid Mars-Like Soils: Implications for Soil Habitability

Soil samples from the hyper-arid region in the Atacama 23 Desert in Southern Peru (La Joya Desert) were analyzed for total and labile organic carbon (TOC & LOC), phospholipid fatty acids analysis (PLFA), quantitative real time polymerase chain reaction (qRT-PCR), 4',6- diamidino-2-phenylindole (DAPI)-fluorescent microscopy, culturable microorganisms, and oxidant activity, in order to understand the relationship between the presence of organic matter and microorganisms in these types of soils. TOC content levels were similar to the labile pool of carbon suggesting the absence of recalcitrant carbon in these soils. The range of LOC was from 2 to 60 micro-g/g of soil. PLFA analysis indicated a maximum of 2.3 x 10(exp 5) cell equivalents/g. Culturing of soil extracts yielded 1.1 x 10(exp 2)-3.7 x 10(exp 3) CFU/g. qRT-PCR showed between 1.0 x 10(exp 2) and 8 x 10(exp 3) cells/g; and DAPI fluorescent staining indicated bacteria counts up to 5 x 104 cells/g. Arid and semiarid samples (controls) showed values between 10(exp 7) and 10(exp 11) cells/g with all of the methods used. Importantly, the concentration of microorganisms in hyper-arid soils did not show any correlation with the organic carbon content; however, there was a significant dependence on the oxidant activity present in these soil samples evaluated as the capacity to decompose sodium formate in 10 hours. We suggest that the analysis of oxidant activity could be a useful indicator of the microbial habitability in hyper-arid soils, obviating the need to measure water activity over time. This approach could be useful in astrobiological studies on other worlds.

Mars-like soils↗

Simple Ion Channels: From Structure to Electrophysiology and Back

A reliable way to establish whether our understanding of a channel is satisfactory is to reproduce its measured ionic conductance over a broad range of applied voltages in computer simulations. In molecular dynamics (MD), this can be done by way of applying an external electric field to the system and counting the number of ions that traverse the channel per unit time. Since this approach is computationally very expensive, we have developed a markedly more efficient alternative in which MD is combined with the electrodiffusion (ED) equation. In this approach, the assumptions of the ED equation can be rigorously tested, and the precision and consistency of the calculated conductance can be determined. We have demonstrated that the full current/voltage dependence and the underlying free energy profile for a simple channel can be reliably calculated from equilibrium or non-equilibrium MD simulations at a single voltage. To carry out MD simulations, a structural model of a channel has to be assumed, which is an important constraint, considering that high-resolution structures are available for only very few simple channels. If the comparison of calculated ionic conductance with electrophysiological data is satisfactory, it greatly increases our confidence that the structure and the function are described sufficiently accurately. We examined the validity of the ED for several channels embedded in phospholipid membranes - four naturally occurring channels: trichotoxin, alamethicin, p7 from hepatitis C virus (HCV) and Vpu from the HIV-1 virus, and a synthetic, hexameric channel, formed by a 21-residue peptide that contains only leucine and serine. All these channels mediate transport of potassium and chloride ions. It was found that the ED equation is satisfactory for these systems. In some of them experimental and calculated electrophysiological properties are in good agreement, whereas in others there are strong indications that the structural models are incorrect.

molecular dynamics (MD↗

Transitory Microbial Habitat in the Hyperarid Atacama Desert

Traces of life are nearly ubiquitous on Earth. However, a central unresolved question is whether these traces always indicate an active microbial community or whether, in extreme environments, such as hyperarid deserts, they instead reflect just dormant or dead cells. Although microbial biomass and diversity decrease with increasing aridity in the Atacama Desert, we provide multiple lines of evidence for the presence of an at times metabolically active, microbial community in one of the driest places on Earth. We base this observation on four major lines of evidence: (i) a physico-chemical characterization of the soil habitability after an exceptional rain event, (ii) identified biomolecules indicative of potentially active cells [e.g., presence of ATP, phospholipid fatty acids (PLFAs), metabolites, and enzymatic activity], (iii) measurements of in situ replication rates of genomes of uncultivated bacteria reconstructed from selected samples, and (iv) microbial community patterns specific to soil parameters and depths. We infer that the microbial populations have undergone selection and adaptation in response to their specific soil microenvironment and in particular to the degree of aridity. Collectively, our results highlight that even the hyperarid Atacama Desert can provide a habitable environment for microorganisms that allows them to become metabolically active following an episodic increase in moisture and that once it decreases, so does the activity of the microbiota. These results have implications for the prospect of life on other planets such as Mars, which has transitioned from an earlier wetter environment to today’s extreme hyperaridity.

Schulze-Makuch, Dirk↗

Emergence of Simple Ion Channels by Self-Assembly of Short Peptides

As protocells transitioned from structures bound by membranes made of simple amphiphiles, such as fatty acids, to structures containing phospholipid membranes, they became nearly impermeable to charged species. Yet, the ability to transport ions across membranes was vital to regulating cellular volume, pH homeostasis, generating energy and sensing the environment. For this reason, life evolved ion channels, protein structures surrounding water-filled pores in the membrane that facilitated ion transport. In contemporary cells, ion channels are complex protein assemblies. Could the same function be carried out, even if less efficiently and selectively, by much simpler peptides at the origins of life? To answer this question, we turned to trichogen (GAIV), an 11-residue peptide that was found to permeabilize membrane, causing leakage of the vesicle content including ions. Since GAIV is synthesized non-gnomically, it represents as excellent model for the earliest, poorly optimized channels. Understanding the mechanism by which short peptides, such as GAIV, mediate ion transport across membranes provides crucial information on the emergence, structure and the mechanism of action of the earliest ion channels. In this study, we used extensive molecular dynamics (MD) simulations to investigate directly spontaneous assembly of GAIV peptide and its structural dynamics, the formation of an induced water pore across the membrane and consequently ion conduction through the membrane.

Wei, Chenyu↗

Ion Channels – From Structure to Electrophysiology and Back

A reliable way to establish whether our understanding of a channel is satisfactory is to reproduce its measured ionic conductance over a broad range of applied voltages in computer simulations. In molecular dynamics (MD), this can be done by way of applying an external electric field to the system and counting the number of ions that traverse the channel per unit time. Since this approach is computationally very expensive, we have developed a markedly more efficient alternative in which MD is combined with the electrodiffusion (ED) equation. In this approach, the assumptions of the ED equation can be rigorously tested, and the precision and consistency of the calculated conductance can be determined. We have demonstrated that the full current/voltage dependence and the underlying free energy profile for a simple channel can be reliably calculated from equilibrium or non-equilibrium MD simulations at a single voltage. Free energy profiles can be obtained from non-equilibrium simulations without a loss of accuracy even without the knowledge of diffusion coefficient. To carry out MD simulations, a structural model of a channel has to be assumed, which is an important constraint, considering that high-resolution structures are available for only very few simple channels. If the comparison of calculated ionic conductance with electrophysiological data is satisfactory, it greatly increases our confidence that the structure and the function are described sufficiently accurately. We examined the validity of the ED for several channels embedded in phospholipid membranes - four naturally occurring channels: trichotoxin, alamethicin, p7 from hepatitis C virus (HCV) and Vpu from the HIV-1 virus, a synthetic, hexameric channel, formed by a 21-residue peptide that contains only leucine and serine and a bacterial pentameric ligand-gated ion channel, GLIC. All these channels mediate transport of potassium and chloride ions. It was found that the ED equation is satisfactory for these systems. In some of them, experimental and calculated electrophysiological properties are in good agreement, whereas in others there are strong indications that the structural models are incorrect.

Pohorille, Andrew↗

Molecular level insight into non-bilayer structure formation in thylakoid membranes: a molecular dynamics study

In oxygenic photosynthetic organisms, the light reactions are performed by protein complexes embedded in the lipid bilayer of thylakoid membranes (TMs). The organization of the bulk lipid molecules into bilayer structures provide optimal conditions for the build-up of the proton motive force (pmf) and its utilization for ATP synthesis. However, the lipid composition of TMs is dominated by the non-bilayer lipid species monogalactosyl diacylglycerol (MGDG), and functional plant TMs, besides the bilayer, contain large amounts of non-bilayer lipid phases. Bulk lipids have been shown to be associated with lumenal, stromal-side and marginal-region proteins and proposed to play roles in the self-assembly and photoprotection of the photosynthetic machinery. Furthermore, it has recently been pointed out that the generation and utilization of pmf for ATP synthesis according to the ‘protet’ or protonic charge transfer model Kell (Biochim Biophys Acta Bioenerg 1865(4):149504, 2024), requires high MGDG content Garab (Physiol Plant 177(2):e70230, 2025). In this study, to gain better insight into the structural and functional roles of MGDG, we employed all atom and coarse-grained molecular dynamics simulations to explore how temperature, hydration levels and varying MGDG concentrations affect the structural and dynamic properties of bilayer membranes constituted of plant thylakoid lipids. Our findings reveal that MGDG promotes increased membrane fluidity and dynamic fluctuations in membrane thickness. MGDG-rich stacked bilayers spontaneously formed inverted hexagonal phases; these transitions were enhanced at low hydration levels and at elevated but physiologically relevant temperatures. It can thus be inferred that MGDG plays important roles in heat and drought stress mechanisms.

59 BASIC BIOLOGICAL SCIENCES↗

Microalgal lipids: biochemistry and biotechnology

Microalgae, including cyanobacteria, are a genetically diverse and biochemically diverse group of mostly photosynthetic organisms that can be found in nearly every ecosystem. They produce a broad range of compounds, including structural and bioactive lipids with various roles in the metabolism of the cell and potential applications in biotechnology. Here, the majority of lipids are synthesized in the chloroplast using acetate to produce fatty acids and terpenoids via the acetate pathway and methylerythritol phosphate deoxy-xylulose phosphate pathway, respectively. This review will highlight the major groups of microalgal lipids as well as their applications in food, fuels, and specialty chemicals. Future directions include the development of microalgal chassis organisms and molecular tools to manipulate lipid synthesis for the enhanced production of target metabolites.

cyanobacteria↗

Identification of structural transitions in bacterial fatty acid binding proteins that permit ligand entry and exit at membranes

Fatty acid (FA) transfer proteins extract FA from membranes and sequester them to facilitate their movement through the cytosol. Detailed structural information is available for these soluble protein–FA complexes, but the structure of the protein conformation responsible for FA exchange at the membrane is unknown. Staphylococcus aureus FakB1 is a prototypical bacterial FA transfer protein that binds palmitate within a narrow, buried tunnel. Here, we define the conformational change from a “closed” FakB1 state to an “open” state that associates with the membrane and provides a path for entry and egress of the FA. Using NMR spectroscopy, we identified a conformationally flexible dynamic region in FakB1, and X-ray crystallography of FakB1 mutants captured the conformation of the open state. In addition, molecular dynamics simulations show that the new amphipathic α-helix formed in the open state inserts below the phosphate plane of the bilayer to create a diffusion channel for the hydrophobic FA tail to access the hydrocarbon core and place the carboxyl group at the phosphate layer. The membrane binding and catalytic properties of site-directed mutants were consistent with the proposed membrane docked structure predicted by our molecular dynamics simulations. Finally, the structure of the bilayer-associated conformation of FakB1 has local similarities with mammalian FA binding proteins and provides a conceptual framework for how these proteins interact with the membrane to create a diffusion channel from the FA location in the bilayer to the protein interior.

37 INORGANIC, ORGANIC, PHYSICAL, AND ANALYTICAL CH↗