共查询到20条相似文献,搜索用时 15 毫秒
1.
Petra C. Gufler 《生物化学与生物物理学报:生物膜》2004,1661(2):154-165
In the present work, S-layer supported lipid membranes formed by a modified Langmuir-Blodgett technique were investigated by electrochemical impedance spectroscopy (EIS). Basically two intermediate hydrophilic supports for phospholipid- (DPhyPC) and bipolar tetraetherlipid- (MPL from Thermoplasma acidophilum) membranes have been applied: First, the S-layer protein SbpA isolated from Bacillus sphaericus CCM 2177 recrystallized onto a gold electrode; and second, as a reference support, an S-layer ultrafiltration membrane (SUM), which consists of a microfiltration membrane (MFM) with deposited S-layer carrying cell wall fragments. The electrochemical properties and the stability of DPhyPC and MPL membranes were found to depend on the used support. The specific capacitances were 0.53 and 0.69 μF/cm2 for DPhyPC bilayers and 0.75 and 0.77 μF/cm2 for MPL monolayers resting on SbpA and SUM, respectively. Membrane resistances of up to 80 MΩ cm2 were observed for DPhyPC bilayers on SbpA. In addition, membranes supported by SbpA exhibited a remarkable long-term robustness of up to 2 days. The membrane functionality could be demonstrated by reconstitution of membrane-active peptides such as valinomycin and alamethicin. The present results recommend S-layer-supported lipid membranes as promising structures for membrane protein-based biosensor technology. 相似文献
2.
生物被膜的形成及其电化学阻抗检测 总被引:1,自引:0,他引:1
生物被膜是细菌及其自身分泌的胞外聚合物组成的微生物群落,其形成是受多种机制共同调控的多阶段动态过程,具有较强的耐药性且难以清除,给医疗、食品等行业带来了巨大的威胁。近年来,生物被膜的相关研究领域备受关注,尤其是针对生物被膜的有效检测技术。本文在简要介绍生物被膜的特点、形成过程及群感效应对生物被膜的调控作用基础之上,总结了生物被膜常用的检测方法,重点针对电化学阻抗技术在生物被膜检测中的应用进行调研和讨论,并对基于微流控芯片的生物被膜电化学阻抗原位检测进行了综述和展望。 相似文献
3.
A. Heyer F. D’Souza X. Zhang G. Ferrari J. M. C. Mol J. H. W. de Wit 《Biodegradation》2014,25(1):67-83
This research paper addresses the biodegradation process for ballast tank coatings in marine environments. As part of this new approach, a commercially available ballast tank coating was exposed to bacteria obtained from a culture collection and to a natural bacterial community isolated from a real ballast tank. The natural community was chosen to explore the interaction of natural biofilms with the coating, an aspect, which is not covered in standard procedures. It is shown that biological activity significantly affects the coating properties. Micro-cracks and holes have been identified using AFM. Acidic bacteria generated holes with 0.2–0.9 μm in depth and 4–9 μm in width. Whereas the natural community additionally caused cracks of 2–8 μm in depth and 1 μm in length. The overall effect of this degradation was examined using the EIS technique. However, the bacterial affected coatings (exposed to acid producing bacteria and a natural community) show a decrease in corrosion resistance. Impedance IZI values decreased over time from 1.18 × 109 to 1.87 × 107 Ω for acidic bacteria and from 1.71 × 109 to 2.24 × 107 Ω for the natural community, indicating a clear loss in coating resistance over time. It is also revealed that the coating corrosion resistance declines after 40 days of exposure for the natural community, leading to the formation of blisters. Bacterial settling could be linked to some specific biofilm patterns affecting different types of coating attack. It can be concluded that it is necessary to include natural communities in coating degradation studies to identify possible degradation mechanisms and the severity of the attack over time. 相似文献
4.
Electrochemical impedance spectroscopy performed on surface-supported bilayer membranes allows for the monitoring of changes in membrane properties, such as thickness, ion permeability, and homogeneity, after exposure to antimicrobial peptides (AMPs). We show that two model cationic peptides, very similar in sequence but different in activity, induce dramatically different changes in membrane properties as probed by impedance spectroscopy. Moreover, the impedance results excluded the “barrel-stave” and the “toroidal pore” models of AMP mode of action, and are more consistent with the “carpet” and the “detergent” models. The impedance data provide important new insights about the kinetics and the scale of the peptide action which currently are not addressed by the “carpet” and the “detergent” models. The method presented not only provides additional information about the mode of action of a particular AMP, but offers a means of characterizing AMP activity in reproducible, well-defined quantitative terms. 相似文献
5.
Radek Macháň 《生物化学与生物物理学报:生物膜》2010,1798(7):1377-2739
Investigation of lipid lateral mobility in biological membranes and their artificial models provides information on membrane dynamics and structure; methods based on optical microscopy are very convenient for such investigations. We focus on fluorescence correlation spectroscopy (FCS), explain its principles and review its state of the art versions such as 2-focus, Z-scan or scanning FCS, which overcome most artefacts of standard FCS (especially those resulting from the need for an external calibration) making it a reliable and versatile method. FCS is also compared to single particle tracking and fluorescence photobleaching recovery and the applicability and the limitations of the methods are briefly reviewed. We discuss several key questions of lateral mobility investigation in planar lipid membranes, namely the influence which membrane and aqueous phase composition (ionic strength and sugar content), choice of a fluorescent tracer molecule, frictional coupling between the two membrane leaflets and between membrane and solid support (in the case of supported membranes) or presence of membrane inhomogeneities has on the lateral mobility of lipids. The recent FCS studies addressing those questions are reviewed and possible explanations of eventual discrepancies are mentioned. 相似文献
6.
7.
This paper reports results of biodegradation studies of polyimide coatings exposed to a mixed fungal culture using electrochemical impedance spectroscopy (EIS). The fungal consortium was originally isolated from degraded polyimides and identified species include Aspergillus versicolor, Cladosporium cladosporioides, and a Chaetomium species. Actively growing fungi on polyimides yield distinctive EIS spectra through time, indicative of failure of the polymer integrity compared to the uninoculated controls. An initial decline in coating resistance was related to the partial ingress of water molecules and ionic species into the polymeric matrices. This was followed by further degradation of the polymers by activity of the fungi. The relationship between the changes in impedance spectra and microbial degradation of the coatings was further supported by scanning electron microscopy, showing extensive colonization of the polyimide surfaces by the fungi. Our data indicate that EIS can be a sensitive and informative technique for evaluating the biosusceptibility of polymers and coatings. 相似文献
8.
The pH-dependent interaction of cinnamomin with lipid membranes investigated by fluorescence methods
Cinnamomin, a new type II ribosome-inactivating protein (RIP), was found to be able to induce the release of calcein loaded in lecithin small unilamellar vesicles and the fusion or aggregation of the lecithin liposomes. Such induction could be promoted severalfold by a pH 5.0 environment, a condition similar to that in endocytic vesicles. Lowering the pH from 7.5 to 5.0 evoked conformational changes of cinnamomin and unmasked its hydrophobic areas, including the exposure of 1-anilino-8-naphthalenesulfonate (1,8-ANS) binding sites of the molecule. Some tryptophan residues with affinity to acrylamide were demonstrated to participate in the lipid-protein interaction. The pH dependent fusogenicity of type II RIP might suggest its in vivo function as a fusogen to exert its cytotoxicity. 相似文献
9.
Binder H 《European biophysics journal : EBJ》2007,36(4-5):265-279
The ordering and H-bonding characteristics of the hydration water of the lipid 1-palmitoyl-2-oleoylphosphatidylcholine (POPC)
were studied using polarized infrared spectroscopy by varying either the temperature or the relative humidity of the ambient
atmosphere of multibilayer samples. The OH-stretching band of lipid-bound water was interpreted by a simplified two-state
model of well-structured, low density “network” water and of less-structured dense “multimer” water. The IR-spectroscopic
data reflect a rather continuous change of the water properties with increasing distance from the membrane and with changing
temperature. Network and multimer water distribute across the whole polar interphase with changing composition and orientation.
Upon dehydration the fraction of network water increases from about 30 to 60%, a value which is similar to that in supercooled
water at −25°C. The highly ordered gel phase gives rise to an increased fraction of structured network water compared with
the liquid crystalline phase. The IR order parameter shows that the water dipoles rearrange from a more parallel towards a
more perpendicular orientation with respect to the membrane normal with progressive hydration.
Dedicated to Prof. K. Arnold on the occasion of his 65th birthday. 相似文献
10.
Plasmon-waveguide resonance and impedance spectroscopy studies of the interaction between penetratin and supported lipid bilayer membranes 总被引:1,自引:0,他引:1
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The interaction between the cell-penetrating peptide, penetratin, and solid-supported lipid bilayer membranes consisting of either egg phosphatidylcholine (PC) or a 75/25 mol% mixture of egg PC and palmitoyloleylphosphatidylglycerol has been studied by simultaneously measuring plasmon-waveguide resonance (PWR) spectra and impedance spectra of lipid-peptide mixtures. When penetratin was incorporated into an egg PC + palmitoyloleylphosphatidylglycerol bilayer, PWR measurements showed a hyperbolic increase in the average refractive index and the refractive index anisotropy, with no change in membrane thickness, over a concentration range between 0 and 2 micro M peptide. In the case of an egg PC bilayer, a biphasic dependence was observed, with a decrease in average refractive index and anisotropy and no thickness change occurring between 0 and 5 micro M peptide, and an increase in membrane thickness occurring between 5 and 15 micro M peptide with no further change in the refractive index parameters. For both membranes, the impedance spectroscopy measurements demonstrated that the electrical resistance was not altered by peptide incorporation, whereas a decrease in membrane capacitance occurred with the same concentration dependence as observed in the PWR experiments, although for the PC membrane no further changes in electrical properties were observed in the higher concentration range. A structural interpretation of these results is described, in which the peptide binds electrostatically within the headgroup region of the bilayer and influences the headgroup conformation, amount of bound water, and the lipid-packing density, without perturbing the hydrocarbon core of the bilayer. 相似文献
11.
The ability to immobilize DNA probes onto gold substrates at an optimum surface density is key in the development of a wide range of DNA biosensors. We present a method to accurately control probe DNA surface density by the simultaneous co-immobilization of thiol modified probes and mercaptohexanol. Probe surface density is controlled by the thiol molar ratio in solution, with a linear relationship between thiol molar ratio and probe density spanning (1-9) x10(12)/cm2. The probe surface density per microscopic surface area was determined using chronocoulometry, and a detailed analysis of the method presented. Using this sample preparation method, the effect of probe density and hybridization on the charge transfer resistance with the negatively charged ferri/ferrocyanide redox couple was determined. Above a threshold probe surface density of 2.5 x 10(12)/cm2, electrostatic repulsion from the negatively charged DNA modulates the charge transfer resistance, allowing hybridization to be detected. Below the threshold density no change in charge transfer resistance with probe density or with hybridization occurs. The probe surface density was optimized to obtain the maximum percentage change in charge transfer resistance with hybridization. 相似文献
12.
Hou Y Helali S Zhang A Jaffrezic-Renault N Martelet C Minic J Gorojankina T Persuy MA Pajot-Augy E Salesse R Bessueille F Samitier J Errachid A Akimov V Reggiani L Pennetta C Alfinito E 《Biosensors & bioelectronics》2006,21(7):1393-1402
Rhodopsin, the G protein-coupled receptor (GPCR) which mediates the sense of vision, was prepared from calf eyes and used as receptor enriched membrane fraction. In this study it was immobilized onto gold electrode by two different techniques: Langmuir-Blodgett (LB) and a strategy based on a self-assembled multilayer. We demonstrated that Langmuir and LB films of rhodopsin are not stable. Thus, in this study a new protein multilayer was prepared on gold electrode by building up layer-by-layer a self-assembled multilayer. It is composed of a mixed self-assembled monolayer formed by MHDA and biotinyl-PE, followed by a biotin-avidin system which allows binding of biotinylated antibody specific to rhodopsin. The immobilization of rhodopsin in membrane fraction, by the specific antibody bound previously on self-assembled multilayer, was monitored with electrochemical impedance spectroscopy (EIS). In addition, the specificity and sensitivity of this self-assembled multilayer system to the presence of rhodopsin were investigated. No effect was observed when the system was in contact with olfactory receptor I7 in membrane fraction used for control measurements. All these results demonstrate that rhodopsin can be immobilized efficiently, specifically, quantitatively and stably on gold electrode through the self-assembled multilayer. 相似文献
13.
Zero mode waveguides (ZMWs), subwavelength optical nanostructures with dimensions ranging from 50 to 200 nm, have been used to study systems involving ligand-receptor interactions. We show that under proper conditions, lipid membranes will invaginate into the nanostructures, which confine optical excitation to subattoliter volumes. Fluorescence correlation spectroscopy (FCS) was used to characterize the diffusion of fluorescently tagged lipids in liquid-disordered phase 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine (POPC) and gel phase 1,2-distearoyl-sn-glycero-3-phosphocholine (DSPC) membranes incubated on the nanostructured surface. In contrast to the POPC, DSPC membranes did not appear to enter the structures, suggesting that invagination is dependent on membrane rigidity. Although correlation curves obtained from POPC membranes conformed to previously derived models for diffusion in the evanescent field within the nanostructure, the diffusion constants obtained were systematically lower than expected. The validity of the one-dimensional diffusion model for membrane diffusion is discussed and it is concluded that the erroneous diffusion constants are a result of nontrivial membrane conformation within the ZMWs. Additionally, FCS was used to characterize the fraction of fluorescently labeled tetanus toxin C fragment bound to a ganglioside-populated POPC membrane within the ZMWs. This allowed the determination of the toxin's equilibrium binding constant at a concentration of 500 nM; higher than possible with diffraction-limited FCS. To our knowledge, the results presented here are the first reported for supported lipid bilayers in nanostructured devices. Furthermore, they open the possibility of studying membrane imbedded receptors and proteins at physiological concentrations with single-molecule resolution. 相似文献
14.
Light scattering by thermal fluctuations on simple monoglyceride bilayer membranes has been used to investigate the viscoelastic properties of these structures. Spectroscopic analysis of these fluctuations (capillary waves) permits the nonperturbative measurement of the interfacial tension and a shear interfacial viscosity acting normal to the membrane plane. The methods were established by studies of solvent and nonsolvent bilayers of glycerol monooleate (GMO). Changes in the tension of GMO/n-decane membranes induced by altering the composition of the parent solution were detected and quantified. In a test of the reliability of the technique controlled variations of the viscosity of the aqueous bathing solution were accurately monitored. The technique was applied to solvent-free bilayers formed from dispersions of GMO in squalane. The lower tensions observed attested to the comparative absence of solvent in such bilayers. In contrast to the solvent case, the solvent-free membranes exhibited a significant transverse shear viscosity, indicative of the enhanced intermolecular interactions within the bilayer. 相似文献
15.
Köhler G Moya SE Leporatti S Bitterlich C Donath E 《European biophysics journal : EBJ》2007,36(4-5):337-347
The interaction between lipid layers supported by polyelectrolyte multilayer cushions has been studied by means of colloidal
force spectroscopy. In a typical experiment, a colloidal probe engineered with a layer-by-layer film and a lipid bilayer on
top is approached to a planar surface coated in a symmetrical way. Kinks of a few nanometres in width appear when lipid layers
are pressed together—reflecting either fusion processes between lipid layers or membranes, or the penetration of polymer blobs
into or through the lipid layers. Retracting curves show a stepwise shape, which results from lipid tether formation or from
polymer stretching, the latter suggesting that polyelectrolyte multilayers make contact as a result of penetration or lipid
fusion.
Dedicated to Prof. K. Arnold on the occasion of his 65th birthday. 相似文献
16.
Lipid membranes were assembled on polyelectrolyte (PE)-coated colloidal particles. The assembly was studied by means of confocal microscopy, flow cytometry, scanning force microscopy, and freeze-fracture electron microscopy. A homogeneous lipid coverage was established within the limits of optical resolution. Flow cytometry showed that the lipid coverage was uniform. Freeze-fracture electron microscopy revealed that the lipid was adsorbed as a bilayer, which closely followed the surface profile of the polyelectrolyte support. Additional adsorption of polyelectrolyte layers on top of the lipid bilayer introduced inhomogeneities as evident from jumps in the fracture plane. Characteristic lipid multilayers have not been seen with freeze-fracture electron microscopy. 相似文献
17.
Bacteriorhodopsin and the nicotinic acetylcholine receptor were biotinylated and reconstituted in lipidic membranes on silicon supports by fusion with proteoliposomes. The presence and distribution of the proteins were studied by binding with streptavidin. Radio-labelled streptavidin was employed for quantifying the amounts of protein remaining in the supported membranes after storage in buffer. The proteins within the membranes remained bound to the surface for weeks. The biological activity of reconstituted unlabelled receptor upon storage showed stability in membranes formed on silicon supports and a reduced stability when formed onto lipid monolayer covered supports. Atomic force microscopy studies on preparations in liquid showed bilayer structures but also attached, partly fused liposomes and membrane particles. In air, the surface was smoother and contained less of liposomes and more of stacked lipid layers. Preparations labelled with streptavidin conjugated to colloidal gold and imaged in air showed the proteins individually distributed, with no protein-rich patches or protein aggregates. 相似文献
18.
The membrane location of the local anesthetics (LA) lidocaine, dibucaine, tetracaine, and procaine hydrochloride as well as their influence on phospholipid bilayers were studied by 31P and 1H magic-angle spinning (MAS) NMR spectroscopy. The 31P NMR spectra of the LA/lipid preparations confirmed that the overall bilayer structure of the membrane remained preserved. The relation between the molecular structure of the LAs and their membrane localization and orientation was investigated quantitatively using induced chemical shifts, nuclear Overhauser enhancement spectroscopy, and paramagnetic relaxation rates. All three methods revealed an average location of the aromatic rings of all LAs in the lipid-water interface of the membrane, with small differences between the individual LAs depending on their molecular properties. While lidocaine is placed in the upper chain/glycerol region of the membrane, for dibucaine and procaine the maximum of the distribution are slightly shifted into the glycerol region. Finally for tetracaine the aromatic ring is placed closest to the aqueous phase in the glycerol/headgroup region of the membrane. The hydrophobic side chains of the LA molecules dibucaine and tetracaine were located deeper in the membrane and showed an orientation towards the hydrocarbon core. In contrast the side chains of lidocaine and procaine are oriented towards the aqueous phase. 相似文献
19.
Kahya N Scherfeld D Bacia K Poolman B Schwille P 《The Journal of biological chemistry》2003,278(30):28109-28115
Confocal fluorescence microscopy and fluorescence correlation spectroscopy (FCS) have been employed to investigate the lipid spatial and dynamic organization in giant unilamellar vesicles (GUVs) prepared from ternary mixtures of dioleoyl-phosphatidylcholine/sphingomyelin/cholesterol. For a certain range of cholesterol concentration, formation of domains with raft-like properties was observed. Strikingly, the lipophilic probe 1,1'-dioctadecyl-3,3,3',3'-tetramethylindocarbocyanine perchlorate (DiI-C18) was excluded from sphingomyelin-enriched regions, where the raft marker ganglioside GM1 was localized. Cholesterol was shown to promote lipid segregation in dioleoyl-phosphatidylcholine-enriched, liquid-disordered, and sphingomyelin-enriched, liquid-ordered phases. Most importantly, the lipid mobility in sphingomyelin-enriched regions significantly increased by increasing the cholesterol concentration. These results pinpoint the key role, played by cholesterol in tuning lipid dynamics in membranes. At cholesterol concentrations >50 mol%, domains vanished and the lipid diffusion slowed down upon further addition of cholesterol. By taking the molecular diffusion coefficients as a fingerprint of membrane phase compositions, FCS is proven to evaluate domain lipid compositions. Moreover, FCS data from ternary and binary mixtures have been used to build a ternary phase diagram, which shows areas of phase coexistence, transition points, and, importantly, how lipid dynamics varies between and within phase regions. 相似文献
20.
Herein, we report an anomalous electrochemical behavior of surface-bound DNA duplex that has single-base mismatches at its distal end. Single-stranded 15-base DNA was immobilized at its 5'end onto gold electrode surfaces. After hybridization with complementary or mismatched DNA, electrochemical impedance spectra were obtained using [Fe(CN)(6)]3-/4- as redox marker ions. Hybridization with the complementary DNA reduced the charge-transfer resistance (R(CT)), whereas single-base mismatches at the distal end of the duplex largely increased the R(CT). This anomaly was found only with the distal end: the increase in R(CT) was not observed for mismatches at either the middle or the proximal end. These results indicate that electrochemical detection of single-base alterations at an end of sample DNA is exceptionally easy because of the diametrically opposite responses. This detection principle is promising for the typing of single-nucleotide polymorphisms in combination with the single-base primer extension protocol. 相似文献