Packing Stress Research Articles

The effects of plant flavones on the membrane boundary potential and lipid packing stress

Here we have revealed the effects of different plant flavones on the physicochemical properties of model lipid membranes. We have demonstrated that baicalein increases the boundary potential of membranes composed of phosphatidylcholine, while wogonin does not affect it. Other flavones tested reduce membrane boundary potential, with this ability increasing among scutellarein, chrysin, apigenin, morin, fisetin, and luteolin. Molecular dynamics simulations demonstrate connection of alteration in boundary potential with the preferential orientation of intrinsic flavone dipole moments in membranes. We have also shown that flavones reduce the melting point of phosphatidylcholine, and this ability increases in the series of luteolin, morin, wogonin, scutellarein, apigenin, baicalein, chrysin, and fisetin. The introduction of baicalein, chrysin and fisetin also leads to a significant decrease in the sharpness of the lipid phase transition. We have hypothesized that the localization of flavones in the glycerol backbone or in the C1-C8 methylene region of lipid hydrocarbon chains leads to an increase in the area per lipid and, as a consequence, to an expansion of the lipid melting peak. Replacement of neutral phosphatidylcholine with negatively charged phosphatidylserine affects the membrane-modifying activity of flavones which given the externalization of phosphatidylserine on the surface of cancer cells may be crucial in the flavone anticancer effects.

Different lateral packing stress in acyl chains alters KcsA orientation and structure in lipid membranes

The molecular structures of the various intrinsic lipids in membranes regulate lipid-protein interactions. These different lipid structures with unique volumes produce different lipid molecular packing stresses/lateral stresses in lipid membranes. Most studies examining lipid packing effects have used phosphatidylcholine and phosphatidylethanolamine (PE), which are the main phospholipids of eukaryotic cell membranes. In contrast, Gram-negative or Gram-positive bacterial membranes are composed primarily of phosphatidylglycerol (PG) and PE, and the physical and thermodynamic properties of each acyl chain in PG at the molecular level remain unresolved. In this study, we used 1-palmitoyl-2-oleoyl-sn-glycero-3-phospho-(1′-rac-glycerol) (POPG, 16:0–18:1 PG) and 1-palmitoyl-2-arachidonoyl-sn-glycero-3-phospho-(1′-rac-glycerol) (PAPG, 16:0–20:4 PG) to prepare lipid bilayers (liposome) with the rod-type fluorescence probe DPH. We measured the lipid packing conditions by determining the rotational freedom of DPH in POPG or PAPG bilayers. Furthermore, we investigated the effect of different monoacyl chains on a K+ channel (KcsA) structure when embedded in POPG or PAPG membranes. The results revealed that differences in the number of double bonds and carbon chain length in the monoacyl chain at sn-2 affected the physicochemical properties of the membrane and the structure and orientation of KcsA.

Toward Predicting the Assembly of Modular Intermetallics from Chemical Pressure Analysis: The Interface Nucleus Approach.

Complex intermetallic phases are often constructed from domains derived from simpler structures arranged into hierarchical assemblies. These modular arrangements offer intriguing prospects, such as the integration of the properties of distinct compounds into a single material or for the emergence of new properties from the interactions among different domains. In this article, we develop a strategy for the design of such complex structures, which we term the interface nucleus approach. Within this framework, the assembly of complex structures is facilitated by interface nuclei: geometrical motifs shared by two parent structures that serve as a region of overlap to nucleate or seed the formation of a combined structure. Our central hypothesis is that the formation of an interface between structures at these motifs creates opportunities for the relief of atomic packing stresses, as revealed by Density Functional Theory-Chemical Pressure (DFT-CP) analysis: when corresponding interatomic contacts in two structures exhibit complementarity─negative CP with positive CP or intense CP with mild CP─the intergrowth allows for a more balanced packing arrangement. To illustrate the application of the interface nucleus concept, we analyze three modular intermetallic structures, the σ-phase (FeCr), PuNi3, and Ca6Cu6Al5 types. In each case, the assembly of the structure can be connected to complementary CP features in an interface nucleus shared by its parent structures, while the distribution of the interface nuclei in the parents serves to template the geometry of the overall framework. In this way, the interface nucleus approach points toward avenues for the design of modular intermetallics from the CP schemes of potential partner structures.

원전용 밸브 패킹응력 해석을 위한 모델링 연구

The valve used in nuclear power plant must be qualified by the known experimental equation, which is the requirement of ASME QME-1 standard. Up to now, the stress on packing is assumed to be constant in design and experimental analysis. This paper deals with the development of analysis model for the packing stress. In the analysis model, the stress is calculated with the love function, whose eigen function is deduced by applying the separation method of axial and radial variables to biharmonic equation with boundary conditions. And the unkown parameters of love function are calculated by applying the remaining boundary conditions. The distribution of radial and axial stress are not constant and the big variation of stress is found at the inner and outer face. The radial stress after boundary layer of radial stress is 45% of gland packing stress. Considering the discussion that the double packing stress makes the radial stress in the packing as same as the fluid pressure, it is proper that the industrial design guide has been used to select the gland packing stress as twice as much as fluid pressure. And the force on stem can be calculated by applying shear stress of packing on stem area. Later, the load factor, which is used in the experimental analysis for consideration of stress distribution, must be related to the real stress distribution by this research result.

From angular to round: in depth interfacial analysis of binary phosphatidylethanolamine mixtures in the inverse hexagonal phase.

Packing stress in the lipidic inverse hexagonal HII phase arises from the necessity of the ideally cylinder-shaped micelles to fill out the hexagonally-shaped Wigner-Seitz unit cell. Thus, hydrocarbon chains stretch towards the corners and compress in the direction of the flat side of the hexagonal unit cell. Additionally, the lipid/water interface deviates from being perfectly circular. To study this packing frustration in greater detail, we have doped 1-palmitoyl-2-oleoyl-sn-phosphatidylethanolamine (POPE) with increasing molar concentrations of 1,2-palmitoyl-sn-phosphatidylethanolamine (DPPE: 0 to 15 mol%). Due to its effectively longer hydrophobic tails, DPPE tends to aggregate in the corner regions of the unit cell, and thus, increases the circularity of the lipid/water interface. From small angle X-ray diffraction (SAXD) we determined electron density maps. Using those, we analysed the size, shape and homogeneity of the lipid/water interface as well as that of the methyl trough region. At 6 and 9 mol% DPPE the nanotubular water core most closely resembles a circle; further to this, in comparison to its neighbouring concentrations, the 9 mol% DPPE sample has the smallest water core area and smallest number of lipids per circumference, best alleviating the packing stress. Finally, a three-water layer model was applied, discerning headgroup, perturbed and free water, demonstrating that the hexagonal phase is most stable in the direction of the flat faces (compression zones) and least stable towards the vertices of the unit cell (decompression zones).

Interplay of Fusion, Leakage, and Electrostatic Lipid Clustering: Membrane Perturbations by a Hydrophobic Antimicrobial Polycation

Membrane active compounds are able to induce various types of membrane perturbations. Natural or biomimetic candidates for antimicrobial treatment or drug delivery scenarios are mostly designed and tested for their ability to induce membrane permeabilization, also termed leakage. Furthermore, the interaction of these usually cationic amphiphiles with negatively charged vesicles often causes colloidal instability leading to vesicle aggregation or/and vesicle fusion. We show the interplay of these modes of membrane perturbation in mixed phosphatidyl glycerol (PG)/phosphatidyl ethanolamine (PE) by the statistical copolymer MM:CO comprising, both, charged and hydrophobic subunits. MM:CO is a representative of partially hydrophobic, highly active, but less selective antimicrobial polycations. Cryo-electron microscopy indicates vesicle fusion rather than vesicle aggregation upon the addition of MM:CO to negatively charged PG/PE (1:1) vesicles. In a combination of fluorescence-based leakage and fusion assays, there is support for membrane permeabilization and pronounced vesicle fusion activity as distinct effects. To this end, membrane fusion and aggregation were prevented by including lipids with polyethylene glycol attached to their head groups (PEG-lipids). The leakage activity of MM:CO is very similar in the absence and presence of PEG-lipids. Vesicle aggregation and fusion however are largely suppressed. This strongly suggests that MM:CO induces leakage by asymmetric packing stress because of hydrophobically driven interactions which could lead to leakage. As a further membrane perturbation effect, MM:CO causes lipid clustering in model vesicles. We address potential artifacts and misinterpretations of experiments characterizing leakage and fusion. Additional to the leakage activity, the pronounced fusogenic activity of the polymer and potentially of many other similar compounds likely has implications for antimicrobial activity and beyond.

A simple mechanistic explanation for Barth syndrome and cardiolipin remodeling.

Barth syndrome is a multisystem disorder caused by an abnormal metabolism of the mitochondrial lipid cardiolipin. In this review, we discuss physical properties, biosynthesis, membrane assembly, and function of cardiolipin. We hypothesize that cardiolipin reduces packing stress in the inner mitochondrial membrane, which arises as a result of protein crowding. According to this hypothesis, patients with Barth syndrome are unable to meet peak energy demands because they fail to concentrate the proteins of oxidative phosphorylation to a high surface density in the inner mitochondrial membrane.

A study on calculation of friction coefficient and packing stress using static diagnosis test for a balanced globe valve in nuclear power plants

A valve assembly used in nuclear power plants must be qualified and supervised. New technical standards such as ASME QME-1 2007 particularly require detailed qualification using experiment and analysis. Particularly, diagnostic tests and engineering studies are required for qualification of ASME QME-1 2007. Among these studies, the research on the measurement of friction coefficient and packing stress is important. The irregular change of packing stress along the stroke distance occurs because of the abnormal phenomenon, which must be found and studied with quantitative methods. Packing stress should be analyzed conservatively through experimentation and analysis. In this study, various formulas were applied to measure and calculate coefficient of friction and packing stress. This study can be used in relation to qualification and supervision of packing materials. And the calculation using static diagnosis test can be used to find the packing frictional force in dynamic diagnosis test with flow pressure in a pipe. This study has made it possible to reliably consider packing frictional force generated in a valve body. And so, it is believed that more margin can be secured when evaluating the capacity of valve actuator by applying the accurate frictional force generated in the valve assembly.

Analysis of the Performance Improvement Effect of Combined Packing for Oil and Gas Well Packer

Packer is a commonly used downhole tool for oil and gas wells. In order to understand the working performance of combined packing, a finite element analysis is performed on the double expansion-compression-expansion combined seal packers to find the effect of combined seal. Taking packers sealed in casing with a diameter of 114.3mm and a wall thickness of 12.7mm as an example, finite element models are established for double expansion and expansion-compression combined seal packers respectively. By applying packer-setting pressure to the finite element models and processing the strain-stress data resulting from finite element analyses, the distribution rules of packing stress and contact pressure between the packing and the inner casing wall for different combined seal packers are obtained. The working performance of the single expansion seal packer and the double expansion and expansion-compression combined seal packers is compared and analyzed. According to the results of finite element analyses, both double expansion combined seal and expansion-compression combined packings can be in effective contact with the casing under a packer-setting pressure of 15MPa, and the contact pressure between the combined seal packing and the inner casing wall is larger than that of a single seal packing. The combined seal packer therefore can actually improve the sealing and packoff effect of the packer. The expansion-compression combined seal has the highest pressure bearing capacity, which is 50% higher than that of the single expansion packer. The lower packing of the double expansion combined packer and the expansion packing of the expansion-compression combined seal packers should be appropriately strengthened and thickened to improve the packing performance of the combined seal packer. The results of contrastive analysis reveal the improvement effect of the sealing and packoff performance of the combined seal packers, which can be used as a reference for the improved design of such packer.

Vitamin E Promotes the Inverse Hexagonal Phase via a Novel Mechanism: Implications for Antioxidant Role.

Vitamin E (α-tocopherol) and a range of other biological compounds have long been known to promote the HII (inverted hexagonal) phase in lipids. Now, it has been well established that purely hydrophobic lipids such as dodecane promote the HII phase by relieving extensive packing stress. They do so by residing deep within the hydrocarbon core. However, we argue from X-ray diffraction data obtained with 1-palmitoyl-2-oleoylphosphatidylcholine (POPE) and 1,2-dioleoylphosphatidylcholine (DOPE) that α-tocopherol promotes the HII phase by a different mechanism. The OH group on the chromanol moiety of α-tocopherol anchors it near the aqueous interface. This restriction combined with the relatively short length of α-tocopherol (as compared to DOPE and POPE) means that α-tocopherol promotes the HII phase by relieving compressive packing stress. This observation offers new insight into the nature of packing stress and lipid biophysics. With the deeper understanding of packing stress offered by our results, we also explore the role that molecular structure plays in the primary function of vitamin E, which is to prevent the oxidation of polyunsaturated membrane lipids.

Bioactive Metabolites of Marine Origin Have Unusual Effects on Model Membrane Systems.

Marine sponges and soft corals have yielded novel compounds with antineoplastic and antimicrobial activities. Their mechanisms of action are poorly understood, and in most cases, little relevant experimental evidence is available on this topic. In the present study, we investigated whether agelasine D (compound 1) and three agelasine analogs (compound 2–4) as well as malonganenone J (compound 5), affect the physical properties of a simple lipid model system, consisting of dioleoylphospahtidylcholine and dioleoylphosphatidylethanolamine. The data indicated that all the tested compounds increased stored curvature elastic stress, and therefore, tend to deform the bilayer which occurs without a reduction in the packing stress of the hexagonal phase. Furthermore, lower concentrations (1%) appear to have a more pronounced effect than higher ones (5–10%). For compounds 4 and 5, this effect is also reflected in phospholipid headgroup mobility assessed using 31P chemical shift anisotropy (CSA) values of the lamellar phases. Among the compounds tested, compound 4 stands out with respect to its effects on the membrane model systems, which matches its efficacy against a broad spectrum of pathogens. Future work that aims to increase the pharmacological usefulness of these compounds could benefit from taking into account the compound effects on the fluid lamellar phase at low concentrations.

Assembly of the complexes of oxidative phosphorylation triggers the remodeling of cardiolipin

Cardiolipin (CL) is a mitochondrial phospholipid with a very specific and functionally important fatty acid composition, generated by tafazzin. However, in vitro tafazzin catalyzes a promiscuous acyl exchange that acquires specificity only in response to perturbations of the physical state of lipids. To identify the process that imposes acyl specificity onto CL remodeling in vivo, we analyzed a series of deletions and knockdowns in Saccharomyces cerevisiae and Drosophila melanogaster, including carriers, membrane homeostasis proteins, fission-fusion proteins, cristae-shape controlling and MICOS proteins, and the complexes I-V. Among those, only the complexes of oxidative phosphorylation (OXPHOS) affected the CL composition. Rather than any specific complex, it was the global impairment of the OXPHOS system that altered CL and at the same time shortened its half-life. The knockdown of OXPHOS expression had the same effect on CL as the knockdown of tafazzin in Drosophila flight muscles, including a change in CL composition and the accumulation of monolyso-CL. Thus, the assembly of OXPHOS complexes induces CL remodeling, which, in turn, leads to CL stabilization. We hypothesize that protein crowding in the OXPHOS system imposes packing stress on the lipid bilayer, which is relieved by CL remodeling to form tightly packed lipid-protein complexes.

Impact of Topology on the Characteristics of Water inside Cubic Lyotropic Liquid Crystalline Systems.

Water molecules present inside the lipid-based cubic liquid crystalline phases are found to play a major role in wide range of applications, such as protein crystallization, virus detection, delivery of drug and biomolecules, etc. In this regard, it is crucial to elucidate static and dynamic properties of the water molecules in the nanochannels and to explore the effect of geometrical topology on the nature of the water inside the different cubic phases. In the present work, we have incorporated two probes, coumarin-343 (C-343) and coumarin-480 (C-480), in two cubic phases with different symmetries, namely gyroid ( Ia3 d) and double diamond ( Pn3 m) with the same water content (22%), to probe the micropolarity, the microviscosity, and the hydration dynamics at different hydrophobic depths in the mesophases. Steady state results estimate the polarity at the lipid-water interface to be similar to that of ethanol, and the polarity near the more hydrophilic parts of the nanochannel resembles that of ethylene glycol. We have also observed a gradient in the microviscosity inside the LLC nanochannels from time-resolved fluorescence anisotropy studies. The hydration dynamics, which play a key role in the numerous applications of the mesophases, have been probed by the time-dependent Stokes shift method of the two probes, revealing the existence of three kinds of dynamics. The difference in the hydration dynamics inside the two mesophases, where the water molecules confined in the Ia3 d phase exhibit a slower dynamics compared to that in Pn3 m, is the prime importance of this work. The underlying reason for this disparity is majorly associated with the differences in the topology of the two structures including the hydrophobic packing stress, the negative interfacial curvature, and the curvature elastic energy of the lipid-water interface. We believe that this kind of correlation between the structural topology of the different cubic LLC mesophases and nature of the water nanochannel will help to boost the applications of the cubic phases in the future.

Vitamin E Promotes the Inverse Hexagonal Phase Providing Insights on Lipid Packing Stresses: Studies by SAXS and 2H NMR

Vitamin E protects membrane lipids from oxidation. How vitamin E partitions within a membrane and whether it preferentially interacts with polyunsaturated lipids are long-standing questions. Polyunsaturated fatty acids (PUFA) preferentially incorporate into phosphatidylethanolamine (PE) lipids, which have a tendency to form the inverse hexagonal (HII) phase. We present small-angle X-ray scattering (SAXS) and solid state 2H NMR data that illustrate vitamin E promotes the transition to HII phase in PE lipids. It has been established that hydrophobic molecules, such as dodecane, promote the HII phase in PE lipids by packing in the hydrophobic interior and so relieving the extensive stress. The hydrophilic hydroxyl head group on vitamin E, in contrast, is anchored near the aqueous surface while the phytanyl chain extends into the hydrophobic region. Based on analysis of our data we then reason that vitamin E promotes the HII phase by a novel mechanism, instead relieving the compressive stress. These observations provide a deeper understanding of lipid packing stress and potentially give insight into a structural role for vitamin E in its function to protect polyunsaturated lipids that are most vulnerable to oxidative attack.

Structural Basis for the Lipid-Mediated Interaction of Tubulin with VDAC Revealed by Neutron Reflectometry

A preponderance of evidence suggests that the mitochondrial outer membrane (MOM) binds dimeric tubulin, which blocks the flux of metabolites through VDAC, the major MOM channel, by inserting its negatively charged, disordered C-terminal tail into the VDAC pore. The discovery of this novel regulatory mechanism of mitochondrial respiration has raised several fundamental questions, including how tubulin binds to the MOM surface, whether its membrane-bound conformation is consistent with the accessibility of its C-termini to the membrane surface, and what role mitochondrial lipids assume in mediating the tubulin-VDAC interaction. Here we report neutron reflectivity (NR) studies on tubulin-coated and VDAC-containing biomimetic membranes. In combination with molecular dynamics simulations, these reveal, first, that dimeric tubulin is anchored to the membrane surface by an amphipathic a-helix of the a-tubulin subunit in an orientation that presents the C-termini of both tubulin subunits to the membrane surface. The structural model is supported by electrophysiological evidence using recombinant tubulin constructs in which C-terminal tails bound to either subunit are individually observed in a VDAC nanopore. Second, we demonstrate using NR that surface-immobilized VDAC can be reconstituted into a complete lipid membrane, thus creating a “protein-tethered bilayer lipid membrane” (ptBLM) over a large sample area. The ptBLM architecture allows dynamic changes of the solution environment, while NR reveals the corresponding structural response of the protein and its lipid environment. This allows us to experimentally observe the effects of pH-induced changes in VDAC's structure on the local lipid environment. Molecular dynamics simulations are in good agreement with NR experiments. We speculate that local curvature or lipid packing stress induced by membrane-embedded VDAC and its structural dynamics serves to localize cytosolic regulators such as tubulin to the vicinity of the channel.

Pycnogenol® supplementation improved the erythrocyte stability of packed donkeys during the late hot-dry season in Zaria, Nigeria

The aim of this study was to determine the effect of pycnogenol® (PYC) on erythrocyte stability of donkeys subjected to packing during the late hot-dry season. Ten donkeys, divided into two groups, served as experimental subjects. P-PYC group (n = 5) was subjected to packing only, while P+PYC group (n = 5) was administered with PYC (10 mg/kg) and subjected to packing. Erythrocytes obtained from each animal in the experimental groups were added into saline solution of different concentrations (0.9, 0.6, 0.4. 0.2, and 0.0 g/100 ml). Erythrocyte osmotic fragility index (EOFI) was evaluated spectrophotometrically. The concentration of isoprostane was also determined. The mean EOFI was significantly (P 0.05) difference in the MCF recorded in both groups, before, after, 3, and 7 days after packing. Percentage erythrocyte stability was significantly higher (P < 0.05) in P+PYC group, especially during, and 7 days after packing with values of 24.3 ± 9.2 and − 23.3 ± 22.8%, respectively when compared with the values of 8.6 ± 3.3 and − 79.7 ± 9.3% obtained in P-PYC group, respectively. Mean isoprostane concentration was significantly (P < 0.05) higher in P-PYC group when compared with the value in P+PYC group, 7 days after packing. In conclusion, environmental and packing stresses decreased stability of the erythrocytes in donkeys; however, PYC may have played an ameliorative role.

Lipid-mediated mode of action of local anesthetics on lipid pores induced by polyenes, peptides and lipopeptides.

The effects of local anesthetics (LAs), namely, lidocaine (LDC), prilocaine (PLC), mepivacaine (MPV), bupivacaine (BPV), procaine (PC), and tetracaine (TTC), on the steady-state transmembrane conductance induced by the cis-side addition of the antifungal polyene macrolide antibiotic, nystatin (NYS), in planar lipid bilayers were studied. The addition of TTC to model membranes comprising DOPC and cholesterol (33 mol%) led to a nearly twenty-fold increase in the steady-state NYS-induced membrane conductance. BPV slightly enhanced the channel-forming activity of polyene. LDC, PLC, MPV, and PC did not affect the NYS-induced transmembrane current. We concluded that the effects of LAs on the channel-forming activity of NYS were in agreement with their effects on the elastic properties of model membranes. The ability of aminoamide LAs to promote calcein leakage from large unilamellar DOPC-vesicles was decreased in the following order: BPV >> LDC ≈ PLC ≈ MPV. LDC, PLC, and MPV produced a graded leakage of fluorescent marker from liposomes, up to 10-13%. A initial sharp jump in fluorescence after the introduction of BPV was attributed to the solubilization of liposomes and the formation of mixed DOPC:BPV-micelles. Differential scanning microcalorimetry (DSC) of large unilamellar DPPC-vesicles showed that the main transition temperature (Tm) is continuously decreased upon increasing concentrations of TTC. A sharp drop in the enthalpy of the transition at higher TTC concentrations indicated a formation of anesthetic/lipid mixed micelles. In contrast to TTC, PC slightly decreased Tm, broadened the DSC signal and did not provoke vesicle-to-micelle transition. Both the calcein leakage and DSC data together with the results of measurements of threshold voltages that are required to cause the lipid bilayer breakdown might indicate an alteration in the curvature lipid packing stress, induced by BPV and TTC. The data presented here lend support to a lipid-mediated mode of LAs action on NYS pores via an alteration in curvature stress near the trans-mouth. Similar results were obtained for several lipid pores, formed by polyene amphotericin B, lipopeptide syringomycin E, and the peptides magainin and melittin. This finding further developed the concept of non-specific regulation of lipid pores by LAs. In conclusion, the combination of nystatin with LAs could be a novel treatment for efficient therapy of superficial and mucosal candidiasis.

Probing the extended lipid anchorage with cytochrome c and liposomes containing diacylphosphatidylglycerol lipids

Experiments investigating the adsorption and desorption of cytochrome c onto and from liposomes containing 50 mol% 1,2-diacylphosphatidylglycerol lipids [10:0, 12:0, 14:0, 16:0, 18:1(Δ9 cis)] with 1,2-dioleoyl-sn-glycero-3-phosphatidylcholine (DOPC) in pH 7.4 buffered solutions of low to moderate ionic strength are reported. Fluorescence experiments show that cytochrome c has a similar adsorption affinity for the five labeled 50 mol% PG liposome systems investigated. Fluorescence recovery experiments reveal the extent of cytochrome c desorption upon the addition of >10× excess of unlabeled 100% 1,2-dioleoyl-sn-glycero-3-phosphatidylglycerol (DOPG) liposomes is dependent on the lipid's acyl chain length. The extent of desorption is also shown to be independent of temperature, albeit over a narrow range. The differences in the extent of cytochrome c desorption from liposomes containing PG lipids with different acyl chain lengths is attributed to the varying contribution of the binding motif involving the extended lipid anchorage in response to lipid packing stress.

Mitochondrial VDAC as an Electromechanical Probe of Lipid-Dependent Membrane Binding of Alpha-Synuclein

Alpha-synuclein (α-syn) is a small, intrinsically disordered cytoplasmic protein highly expressed in the central nervous system and implicated in Parkinson disease. α-Syn was found to specifically bind to native mitochondrial membranes in live cells and is known to be involved in neurodegenerative mitochondrial dysfunction and in neuroapoptosis. Thus, understanding the molecular mechanisms of α-syn interaction with model membranes mimicking the mitochondrial outer membrane (MOM) is of particular importance. Here we study the α-syn-membrane interaction on a single-molecule level using the voltage dependent anion channel (VDAC) to probe the kinetic and structural rearrangements of α-syn at the lipid membrane. We find that both the frequency and duration of the VDAC blockage by α-syn depend strongly on lipid composition. The highest interaction frequency of α-syn with VDAC is observed in anionic lipids; it drops by 100-fold in cationic lipids. The interaction frequency also increases with nonlamellar phosphatidylethanolamine content in neutral membranes. The duration of blockage is also affected by lipid composition, particularly the onset of α-syn translocation through the VDAC pore. These results support a model in which α-syn's N-terminus binding to the membrane is followed by C-terminal blockage of the VDAC pore. Comparison of electrophysiological data with macroscopic measurements of α-syn-lipid binding using Fluorescence Correlation Spectroscopy and Bilayer Overtone Analysis methods suggests that three domains of α-syn play different roles in membrane interactions and, consequently, with VDAC pore. Our results establish VDAC as a suitable electromechanical probe to report on monomeric α-syn membrane binding kinetics and dynamic conformational adaptations to the lipid charges and lipid packing stress. This system will facilitate further exploration of the well-recognized, but not well understood, regulatory roles of mitochondrial lipids in bioenergetics and cell metabolism.

ILQINS Hexapeptide, Identified in Lysozyme Left-Handed Helical Ribbons and Nanotubes, Forms Right-Handed Helical Ribbons and Crystals

Amyloid fibrils are implicated in over 20 neurodegenerative diseases. The mechanisms of fibril structuring and formation are not only of medical and biological importance but are also relevant for material science and nanotechnologies due to the unique structural and physical properties of amyloids. We previously found that hen egg white lysozyme, homologous to the disease-related human lysozyme, can form left-handed giant ribbons, closing into nanotubes. By using matrix-assisted laser desorption ionization mass spectrometry analysis, we here identify a key component of such structures: the ILQINS hexapeptide. By combining atomic force microscopy and circular dichorism, we find that this fragment, synthesized by solid-phase peptide synthesis, also forms fibrillar structures in water at pH 2. However, all fibrillar structures formed possess an unexpected right-handed twist, a rare chirality within the corpus of amyloid experimental observations. We confirm by small- and wide-angle X-ray scattering and molecular dynamics simulations that these fibrils are composed of conventional left-handed β-sheets, but that packing stresses between adjacent sheets create this twist of unusual handedness. We also show that the right-handed fibrils represent a metastable state toward β-sheet-based microcrystals formation.