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1.
Small ; 17(46): e2102975, 2021 11.
Artículo en Inglés | MEDLINE | ID: mdl-34643032

RESUMEN

Filtration through membranes with nanopores is typically associated with high transmembrane pressures and high energy consumption. This problem can be addressed by reducing the respective membrane thickness. Here, a simple procedure is described to prepare ultrathin membranes based on protein nanopores, which exhibit excellent water permeance, two orders of magnitude superior to comparable, industrially applied membranes. Furthermore, incorporation of either closed or open protein nanopores allows tailoring the membrane's ion permeability. To form such membranes, the transmembrane protein ferric hydroxamate uptake protein component A (FhuA) or its open-pore variant are assembled at the air-water interface of a Langmuir trough, compressed to a dense film, crosslinked by glutaraldehyde, and transferred to various support materials. This approach allows to prepare monolayer or multilayer membranes with a very high density of protein nanopores. Freestanding membranes covering holes up to 5 µm in diameter are visualized by atomic force microscopy (AFM), helium ion microscopy, and transmission electron microscopy. AFM PeakForce quantitative nanomechanical property mapping (PeakForce QNM)  demonstrates remarkable mechanical stability and elastic properties of freestanding monolayer membranes with a thickness of only 5 nm. The new protein membrane can pave the way to energy-efficient nanofiltration.


Asunto(s)
Nanoporos , Membranas Artificiales , Microscopía de Fuerza Atómica , Microscopía Electrónica de Transmisión
2.
Langmuir ; 35(13): 4613-4625, 2019 04 02.
Artículo en Inglés | MEDLINE | ID: mdl-30840475

RESUMEN

In continuation of previous work, we present a new promising DNA carrier, OO4, a highly effective peptide-mimicking lysine-based cationic lipid. The structural characteristics of the polynucleotide carrier system OO4 mixed with the commonly used co-lipid DOPE and the saturated phospholipid DPPE have been studied in two-dimensional and three-dimensional model systems to understand their influence on the physical-chemical properties. The phase behavior of pure OO4 and its mixtures with DOPE and DPPE was studied at the air-water interface using a Langmuir film balance combined with infrared reflection-absorption spectroscopy. In bulk, the self-assembling structures in the presence and absence of DNA were determined by small-angle and wide-angle X-ray scattering. The amount of adsorbed DNA to cationic lipid bilayers was measured using a quartz crystal microbalance. The choice of the co-lipid has an enormous influence on the structure and capability of binding DNA. DOPE promotes the formation of nonlamellar lipoplexes (cubic and hexagonal structures), whereas DPPE promotes the formation of lamellar lipoplexes. The correlation of the observed structures with the transfection efficiency and serum stability indicates that OO4/DOPE 1:3 lipoplexes with a DNA-containing cubic phase encapsulated in multilamellar structures seem to be most promising.


Asunto(s)
ADN/química , Liposomas/química , Cationes/química , Membrana Dobles de Lípidos/química , Fosfolípidos/química
3.
Molecules ; 24(19)2019 Oct 05.
Artículo en Inglés | MEDLINE | ID: mdl-31590402

RESUMEN

Due to the characteristic chain rigidity and weak intermolecular interactions of perfluorinated substances, the phase diagram of Langmuir monolayer formed by perfluorinated molecules has been interpreted so far as displaying only two phases, a 2D gas (G) and a liquid condensed (LC). However, in this work, we presented Grazing Incidence X-ray Diffraction measurements, which exhibit two diffraction peaks on the transition plateau: One is the signature of the hexagonal structure of the LC phase, the second one is associated to the low-density fluid phase and is thus more ordered than expected for a 2D gas or a typical fluid phase. Atomistic molecular dynamics simulations, performed on the transition plateau, revealed the existence of clusters in which domains of vertical molecules organized in a hexagonal lattice coexist with domains of parallel lines formed by tilted molecules, a new structure that could be described as a "2D smectic C" phase. Moreover, the diffraction spectrum calculated from the simulation trajectories compared favorably with the experimental spectra, fully validating the simulations and the proposed interpretation. The results were also in agreement with the thermodynamic analysis of the fluid phase and X-ray Reflectivity experiments performed before and after the transition between these two phases.


Asunto(s)
Ácidos Grasos/química , Tensoactivos/química , Adsorción , Halogenación , Simulación de Dinámica Molecular , Transición de Fase , Termodinámica , Difracción de Rayos X
4.
Phys Chem Chem Phys ; 20(10): 6936-6944, 2018 Mar 07.
Artículo en Inglés | MEDLINE | ID: mdl-29464262

RESUMEN

The influence of the chain composition on the physical-chemical properties will be discussed for five transfection lipids containing the same lysine-based head group. For this purpose, the chain composition will be gradually varied from saturated tetradecyl (C14:0) and hexadecyl (C16:0) chains to longer but unsaturated oleyl (C18:1) chains with double bonds in the cis configuration. In this work, we investigated the lipids as Langmuir monolayers at the air-water-interface in the absence and presence of calf thymus DNA applying different techniques such as infrared reflection absorption spectroscopy (IRRAS) and grazing incidence X-ray diffraction (GIXD). The replacement of saturated tetradecyl (C14:0) and hexadecyl (C16:0) chains by unsaturated oleyl (C18:1) chains increases the fluidity of the lipid monolayer: TH10 < TT10 < OH10 < OT10 < OO10 resulting in a smaller packing density. TH10 forms the stiffest and OO10 the most fluid monolayer in this structure-property study. OO10 has a higher protonation degree compared to the saturated lipids TT10 and TH10 as well as to the hybrids OT10 and OH10 because of a better accessibility of the amine groups. Depending on the bulk pH, different scenarios of DNA coupling to the lipid monolayers have been proposed.


Asunto(s)
Aminas/química , Lípidos/química , Lisina/química , Animales , Bovinos , ADN/química , Interacciones Hidrofóbicas e Hidrofílicas , Estructura Molecular , Propiedades de Superficie , Transfección , Agua/química
5.
Phys Chem Chem Phys ; 20(25): 17393-17405, 2018 Jun 27.
Artículo en Inglés | MEDLINE | ID: mdl-29911233

RESUMEN

Based on previous work, the influence of the chain composition on the physical-chemical properties of five new transfection lipids (TH10, TT10, OH10, OT10 and OO10) containing the same lysine-based head group has been investigated in aqueous dispersions. For this purpose, the chain composition has been gradually varied from saturated tetradecyl (T, C14:0) and hexadecyl (H, C16:0) chains to longer but unsaturated oleyl (O, C18:1) chains with double bonds in the cis configuration. In this work, the lipid dispersions have been investigated in the absence and presence of the helper lipid DOPE and calf thymus DNA by small-angle and wide-angle X-ray scattering (SAXS/WAXS) supplemented by differential scanning calorimetry (DSC), attenuated total reflection Fourier-transform infrared spectroscopy (ATR-FTIR) and Fourier-transform Raman spectroscopy (FTRS). Lamellar and inverted hexagonal mesophases have been observed in single-component systems. In the binary mixtures, the aggregation behaviour changes with an increasing amount of DOPE from lamellar to cubic. The lipid mixtures with DNA show a panoply of mesophases. Interestingly, TT10 and OT10 form cubic lipoplexes, whereas OO10 complexes the DNA sandwich-like between lipid bilayers in a lamellar lipoplex. Surprisingly, the latter is the most effective lipoplex.


Asunto(s)
ADN/química , Membrana Dobles de Lípidos/química , Liposomas/química , Lisina/química , Transfección/métodos , Células A549 , Animales , ADN/metabolismo , Expresión Génica , Proteínas Fluorescentes Verdes/genética , Células HeLa , Humanos , Células LLC-PK1 , Porcinos , Termodinámica
6.
Phys Chem Chem Phys ; 19(30): 20271-20280, 2017 Aug 02.
Artículo en Inglés | MEDLINE | ID: mdl-28726909

RESUMEN

Cationic lipids are considered as non-viral carriers for genetic material used in gene therapy. They have no carcinogenic potential and cause low immune response compared to existing viral systems. The protonation degree of these cationic lipids is a crucial parameter for the binding behavior of polynucleotides (e.g., DNA). Newly synthesized peptide-mimic lysine-based amino-functionalized lipids have been investigated in 2D models as monolayers at the air-liquid interface. Standard surface pressure - area isotherms have been measured to prove the layer stability. Total reflection X-ray fluorescence (TRXF) has been used as a surface sensitive analytical method to estimate the amount of counterions at the head groups. Using a standard sample as a reference, the protonation degree of these cationic lipids can be quantified on buffers with different pH values. It is found that the protonation degree depends linearly on the packing density of the lipid monolayer.


Asunto(s)
Lípidos/química , Lisina/química , Transfección/métodos , Aire , Cationes/química , ADN/química , ADN/metabolismo , Protones , Espectrometría por Rayos X , Agua/química
7.
Langmuir ; 32(19): 4896-903, 2016 05 17.
Artículo en Inglés | MEDLINE | ID: mdl-27142706

RESUMEN

The artificial phospholipid Pad-PC-Pad was analyzed in 2D (monolayers at the air/water interface) and 3D (aqueous lipid dispersions) systems. In the gel phase, the two leaflets of a Pad-PC-Pad bilayer interdigitate completely, and the hydrophobic bilayer region has a thickness comparable to the length of a single phospholipid acyl chain. This leads to a stiff membrane with no spontaneous curvature. Forced into a vesicular structure, Pad-PC-Pad has faceted geometry, and in its extreme form, tetrahedral vesicles were found as predicted a decade ago. Above the main transition temperature, a noninterdigitated Lα phase with fluid chains has been observed. The addition of cholesterol leads to a slight decrease of the main transition temperature and a gradual decrease in the transition enthalpy until the transition vanishes at 40 mol % cholesterol in the mixture. Additionally, cholesterol pulls the chains apart, and a noninterdigitated gel phase is observed. In monolayers, cholesterol has an ordering effect on liquid-expanded phases and disorders condensed phases. The wavenumbers of the methylene stretching vibration indicate the formation of a liquid-ordered phase in mixtures with 40 mol % cholesterol.


Asunto(s)
Colesterol/química , Aire , Colesterol/metabolismo , Modelos Moleculares , Conformación Molecular , Presión , Propiedades de Superficie , Agua/química
8.
Soft Matter ; 12(27): 5854-66, 2016 Jul 06.
Artículo en Inglés | MEDLINE | ID: mdl-27328379

RESUMEN

As a continuation of previous work, the self-assembly process of cationic lipid formulations in the presence and absence of DNA was investigated with respect to binary lipid mixtures suitable as polynucleotide carrier systems. The lipid blends consist of one malonic-acid-based cationic lipid with a varying alkyl chain pattern, either N-{6-amino-1-[N-(9Z)-octadec-9-enylamino]-1-oxohexan-(2S)-2-yl}-N'-{2-[N,N-bis(2-aminoethyl)amino]ethyl}-2-hexadecylpropandiamide () or N-[6-amino-1-oxo-1-(N-tetradecylamino)hexan-(2S)-2-yl]-N'-{2-[N,N-bis(2-aminoethyl)amino]ethyl}-2-hexadecylpropandiamide (), and one neutral co-lipid, either 1,2-di-[(9Z)-octadec-9-enoyl]-sn-glycero-3-phosphocholine (DOPE) or 1,2-di-(hexadecanoyl)-sn-glycero-3-phosphocholine (DPPC). Although the cationic lipids exhibit only slight differences in their structure, the DNA transfer efficiency varies drastically. Therefore, self-assembly was studied in 3D systems by small- and wide-angle X-ray scattering (SAXS and WAXS) and transmission electron microscopy (TEM) as well as in 2D systems by infrared reflection-absorption spectroscopy (IRRAS) on Langmuir films. The investigated lipid mixtures show quite different self-assembly in the absence of DNA, with varying structures from vesicles (/DOPE; /DOPE) and tubes (/DOPE) to discoid structures (/DPPC; /DPPC). Twisted ribbons and sheets, which were stabilized due to hydrogen-bond networks, were found in all investigated lipid mixtures in the absence of DNA. The addition of DNA leads to the formation of lamellar lipoplexes for all the investigated lipid compositions. The lipoplexes differ in crucial parameters, such as the lamellar repeat distance and the spacing between the DNA strands, indicating differences in the binding strength between DNA and the lipid composition. The formation of associates with an ideal charge density might emerge as a key parameter for efficient DNA transfer. Furthermore, the structures observed for the different lipid compositions in the absence of DNA prepare the way for other applications besides gene therapy.


Asunto(s)
Malonatos/química , Fosfolípidos/química , Cationes , ADN/química , Diamida , Liposomas , Dispersión de Radiación , Transfección , Difracción de Rayos X
9.
Chem Phys Lipids ; 198: 28-38, 2016 Jun.
Artículo en Inglés | MEDLINE | ID: mdl-27134131

RESUMEN

Amyloid formation is related to neurodegenerative diseases like Alzheimer's disease or Parkinson's disease. In the molecular onset of the diseases, soluble peptides adopt conformations that are rich in ß-sheet and ultimately form aggregates. How this process is triggered or influenced by membrane binding, or how the membrane integrity is disturbed by the peptide binding and conformational transition is still under debate. In the present study, we systematically examine the effects of ß-sheet prone model peptides on zwitterionic and negatively charged lipids in both mono- and bilayers and in various lipid phase states by infrared reflection absorption spectroscopy, grazing incidence X-ray diffraction, and small and wide angle X-ray scattering. No difference in the interaction of the peptides with zwitterionic or negatively charged lipids was observed. Furthermore, the interaction of ß-sheet prone model peptides leaves the lipid structure largely unaffected. However, the lipid phase state decides upon the mode of interaction. Peptides insert into liquid-expanded layers and interact only with the head groups of liquid-condensed lipid layers. Using a zoo of complementary techniques and critically examining preparation procedures we are able to obtain an unambiguous picture of peptide binding to membranes.


Asunto(s)
Amiloide/química , Amiloide/metabolismo , Membrana Dobles de Lípidos/química , Membrana Dobles de Lípidos/metabolismo , Modelos Moleculares , Secuencia de Aminoácidos , Interacciones Hidrofóbicas e Hidrofílicas , Unión Proteica , Conformación Proteica en Lámina beta , Propiedades de Superficie , Temperatura
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