Langmuir-Blodgett film of hydrophobin protein from Pleurotus ostreatus at the air-water interface

We present results concerning the formation of Langmuir-Blodgett (LB) films of a class I hydrophobin from Pleurotus ostreatus at the air-water interface, and their structure as Langmuir-Blodgett (LB) films when deposited on silicon substrates. LB films of the hydrophobin were investigated by atomic force microscopy (AFM). We observed that the compressed film at the air-water interface exhibits a molecular depletion even at low surface pressure. In order to estimate the surface molecular concentration, we fit the experimental isotherm with Volmer's equation describing the equation of state for molecular monolayers. We found that about (1)/ 10 of the molecules contribute to the surface film formation. When transferred on silicon substrates, compact and uniform monomolecular layers about 2.5 nm thick, comparable to a typical molecular size, were observed. The monolayers coexist with protein aggregates, under the typical rodlet form with a uniform thickness of about 5.0 nm. The observed rodlets appear to be a hydrophilic bilayer and can then be responsible for the surface molecular depletion.     

Surface properties of class ii hydrophobins from Trichoderma reesei and influence on bubble stability

We report the remarkable surface behavior of class II hydrophobin proteins HFBI and HFBII from Trichoderma reesei and the resulting effect that these proteins have on the stability of air bubbles to the process of disproportionation. The surface properties were studied using surface tensiometry and surface shear rheology. Surface tensiometry data show that hydrophobins are very surface active proteins, reducing the surface tension to approximately 30 mN m-1. The rate at which the hydrophobins adsorb at the surface may also be related to the self-assembly behavior in aqueous solution. We further show that hydrophobins form air/water surfaces with high elasticity, the magnitude of which is well in excess of that of surface layers formed by other common proteins used as foam or emulsion stabilizers. The measured surface properties translate to the stability of bubbles with adsorbed hydrophobin, and in this study, we demonstrate the ability of hydrophobin to have a dramatic effect on the rate of disproportionation in some simple bubble dissolution studies.     

Layer thickness of hydrophobin films leads to oscillation in wettability

In nanobiotechnology, the properties of surfaces are often key to sensor applications. If analytes possess a low tolerance or affinity regarding the sensory substrate (surface), then the setup of mediators may be indicated. Hydrophobins enable biocompatible surface functionalization without significant restrictions of the physicochemical substrate properties. Because of the imperfect formation of hydrophobin films, a high variation in surface properties is observed. In this study, we report on the relation between the film thickness of hydrophobin-coated solid surfaces and their wettability. We found that the wettability of protein-coated surfaces strictly depends on the amount of adsorbed protein, as reflected in an oscillation of the contact angles of hydrophobin-coated silicon wafers. Fusion proteins of Ccg2 and HFBI, representatives of class I and II hydrophobins, document the influence of fused peptide tags on the wettability. The orientation of the first crystal nuclei plays a decisive role in the formation of the growing hydrophobin layers. Here, a simple method of deducing the film thickness of hydrophobin assemblies on solid surfaces is presented. The determination of the static contact angle allows the prediction of which part of the protein is exposed to possible analytes.     

静电力驱动蛋白质在疏水蛋白表面的吸附

疏水蛋白是丝状真菌产生的一种外泌蛋白质, 它们可以在不同表面形成双亲性蛋白膜. 疏水蛋白也是一种优良的蛋白质固定化基质, 然而蛋白质在疏水蛋白表面吸附的驱动机制却是未知的. 本文系统研究了不同pH和离子浓度下蛋白质在疏水蛋白表面的吸附. 首先, 用石英晶体微天平技术研究了不同pH和离子浓度下, Ⅰ型疏水蛋白HGFI和Ⅱ型疏水蛋白HFBI在聚苯乙烯表面的吸附. 结果发现, pH和离子强度对HGFI在聚苯乙烯表面的吸附影响较大, 对HFBI的吸附影响与HGFI相比则较小; HGFI在聚苯乙烯表面主要形成的是弹性膜, 而HFBI在聚苯乙烯表面主要形成的是刚性膜. 随后又研究了不同pH和离子浓度下牛血清白蛋白(BSA)和亲和素(Avidin)在HGFI和HFB上吸附, 结果表明, pH和离子强度对BSA和Avidin在HGFI和HFB上吸附有显著影响, 说明BSA和Avidin在两种疏水蛋白上吸附的主要驱动力为静电力. 本文研究结果为实现疏水蛋白表面可控地固定蛋白质提供了理论指导.     

Self-assembled bilayers from the protein HFBII hydrophobin: nature of the adhesion energy

The hydrophobins are a class of amphiphilic proteins which spontaneously adsorb at the air/water interface and form elastic membranes of high mechanical strength as compared to other proteins. The mechanism of hydrophobin adhesion is of interest for fungal biology and for various applications in electronics, medicine, and food industry. We established that the drainage of free foam films formed from HFBII hydrophobin solutions ends with the appearance of a 6 nm thick film, which consists of two layers of protein molecules, that is, it is a self-assembled bilayer (S-bilayer), with hydrophilic domains pointing inward and hydrophobic domains pointing outward. Its formation is accompanied by a considerable energy gain, which is much greater than that typically observed with free liquid films. The experiments at different pH show that this attraction between the "hydrophilic" parts of the HFBII molecules is dominated by the short-range hydrophobic interaction rather than by the patch-charge electrostatic attraction.     

Self-assembly of class II hydrophobins on polar surfaces

Hydrophobins are structural proteins produced by filamentous fungi that are amphiphilic and function through self-assembling into structures such as membranes. They have diverse roles in the growth and development of fungi, for example in adhesion to substrates, for reducing surface tension to allow aerial growth, in forming protective coatings on spores and other structures. Hydrophobin membranes at the air-water interface and on hydrophobic solids are well studied, but understanding how hydrophobins can bind to a polar surface to make it more hydrophobic has remained unresolved. Here we have studied different class II hydrophobins for their ability to bind to polar surfaces that were immersed in buffer solution. We show here that the binding under some conditions results in a significant increase of water contact angle (WCA) on some surfaces. The highest contact angles were obtained on cationic surfaces where the hydrophobin HFBI has an average WCA of 62.6° at pH 9.0, HFBII an average of 69.0° at pH 8.0, and HFBIII had an average WCA of 61.9° at pH 8.0. The binding of the hydrophobins to the positively charged surface was shown to depend on both pH and ionic strength. The results are significant for understanding the mechanism for formation of structures such as the surface of mycelia or fungal spore coatings as well as for possible technical applications.     

Physicochemical study of laminin-related peptides

Four peptide analogues related to the active sequence YIGSR of laminin have been synthesised. The synthesis and chemical characterisation of the peptides are described. Physicochemical properties of these peptides such as surface activity, spreadability, monolayer formation, as well as their interaction with lipid monolayers and bilayers, have been studied by using Langmuir-Blodgett films and liposomes as membrane models. In spite of their good water solubility, these peptides are able to form stable monolayers at the air/water interface and to insert into lipid monolayers. The interaction with bilayers is soft; they are not able to induce the leakage of entrapped CF nor to modify the microviscosity of bilayers in general. Thus in these models electrostatic forces apparently do not play an important role, as we expected previously according to the electrical charge of bilayers, markers and peptides.     

Hydrophobin purification based on the theory of CO2-nanobubbles

Purification is a critical step to obtain hydrophobin HFBII for use in positive applications. In this study, hydrophobin HFBII was produced by Trichoderma reesei via submerged fermentation. Using the CO₂-foam fractionation method yielded a fourfold increase in protein concentration. The foamate (α_L-HFBII) was dried using a nano spray-dryer under optimal temperature. The gushing activity of the dried foamate (α_S-HFBII) decreased. Addition of Tween 80 to the foamate before the drying process partially prevented the deactivation of hydrophobin HFBII. The purity of the powder was enhanced based on the theory of CO₂-nanobubbles in a CO₂-rich environment. The collected CO₂-nanobubbles were added to an apolar–polar system and the interface of these two phases was collected. After evaporation of the apolar phase, the purity of the hydrophobins assembled on the surface of the liquid was significantly improved.     

Relationships between equilibrium spreading pressure and phase equilibria of phospholipid bilayers and monolayers at the air-water interface

The intricate interplay between the bilayer and monolayer properties of phosphatidylcholine (PC), phosphatidylglycerol (PG), and phosphatidylethanolamine (PE) phospholipids, in relation to their polar headgroup properties, and the effects of chain permutations on those polar headgroup properties have been demonstrated for the first time with a set of time-independent bilayer-monolayer equilibria studies. Bilayer and monolayer phase behavior for PE is quite different than that observed for PC and PG. This difference is attributed to the characteristic biophysical PE polar headgroup property of favorable intermolecular hydrogen-bonding and electrostatic interactions in both the bilayer and monolayer states. This characteristic hydrogen-bonding ability of the PE polar headgroup is reflected in the condensed nature of PE monolayers and a decrease in equilibrium monolayer collapse pressure at temperatures below the monolayer critical temperature, T(c) (whether above or below the monolayer triple point temperature, T(t)). This interesting phenomena is compared to equilibrated PC and PG monolayers which collapse to form bilayers at 45 mN/m at temperatures both above and below monolayer T(c). Additionally, it has been demonstrated by measurements of the equilibrium spreading pressure, pie, that at temperatures above the bilayer main gel-to-liquid-crystalline phase-transition temperature, T(m), all liquid-crystalline phospholipid bilayers spread to form monolayers with pie around 45 mN/m, and spread liquid-expanded equilibrated monolayers collapse at 45 mN/m to form their respective thermodynamically stable liquid-crystalline bilayers. At temperatures below bilayer T(m), PC and PG gel bilayers exhibit a drop in bilayer pi(e) values < or =0.2 mN/m forming gaseous monolayers, whereas the value of pic of spread monolayers remains around 45 mN/m. This suggests that spread equilibrated PC and PG monolayers collapse to a metastable liquid-crystalline bilayer structure at temperatures below bilayer T(m) (where the thermodynamically stable bilayer liquid-crystalline phase does not exist) and with a surface pressure of 45 mN/m, a surface chemical property characteristically observed at temperatures above bilayer T(m) (monolayer T(c)). In contrast, PE gel bilayers, which exist at temperatures below bilayer T(m) but above bilayer T(s) (bilayer crystal-to-gel phase-transition temperature), exhibit gel bilayer spreading to form equilibrated monolayers with intermediate pie values in the range of 30-40 mN/m; however, bilayer pie and monolayer pic values remain equal in value to one another. Contrastingly, at temperatures below bilayer T(s), PE crystalline bilayers exhibit bilayer pie values < or =0.2 mN/m forming equilibrated gaseous monolayers, whereas spread monolayers collapse at a value of pic remaining around 30 mN/m, indicative of metastable gel bilayer formation.     

Interactions in lipid stabilised foam films

The interaction between lipid bilayers in water has been intensively studied over the last decades. Osmotic stress was applied to evaluate the forces between two approaching lipid bilayers in aqueous solution. The force–distance relation between lipid mono- or bilayers deposited on mica sheets using a surface force apparatus (SFA) was also measured. Lipid stabilised foam films offer another possibility to study the interactions between lipid monolayers. These films can be prepared comparatively easy with very good reproducibility. Foam films consist usually of two adsorbed surfactant monolayers separated by a layer of the aqueous solution from which the film is created. Their thickness can be conveniently measured using microinterferometric techniques. Studies with foam films deliver valuable information on the interactions between lipid membranes and especially their stability and permeability. Presenting inverse black lipid membrane (BLM) foam films supply information about the properties of the lipid self-organisation in bilayers. The present paper summarises results on microscopic lipid stabilised foam films by measuring their thickness and contact angle. Most of the presented results concern foam films prepared from dispersions of the zwitterionic lipid 1,2-dimyristoyl-sn-glycero-3-phosphorylcholine (DMPC) and some of its mixtures with the anionic lipid — 1,2-dimyristoyl-sn-glycero-3-[phospho-rac-(1-glycerol)] (DMPG).     

Surface modification using a novel type I hydrophobin HGFI

Surface wettability conversion with hydrophobins is important for its applications in biodevices. In this work, the application of a type I hydrophobin HGFI in surface wettability conversion on mica, glass, and poly(dimethylsiloxane) (PDMS) was investigated. X-ray photoelectron spectroscopy (XPS) and water-contact-angle (WCA) measurements indicated that HGFI modification could efficiently change the surface wettability. Data also showed that self-assembled HGFI had better stability than type II hydrophobin HFBI. Protein patterning and the following immunoassay illustrated that surface modification with HGFI should be a feasible strategy for biosensor device fabrication. Figure A hydrophobin HGFI has been applied into surface wettability conversion for protein immobilization Electronic supplementary material  The online version of this article (doi:) contains supplementary material, which is available to authorized users.     

Two-dimensional arrays of amphiphilic Zn2+-cyclens for guided molecular recognition at interfaces

The sterically guided molecular recognition of nucleobases, phosphates, adenosine, and uridine nucleotides on Langmuir monolayers and Langmuir-Blodgett monolayers of amphiphilic mono- or bis(Zn2+-cyclen)s assembled on thiolated surfaces was investigated. The stepwise selective binding of metal ions, uracil, or phosphate by dicetyl cyclen monolayers with variously tuned structures at the air/water interface was corroborated by the measurements of the corresponding LB films deposited onto quartz crystals. Two types of recognition surfaces were fabricated from Zn2+-dicetyl cyclen. The surface covered with a complex preformed in the Langmuir monolayer was capable both of imide and of phosphate binding. The similar complex formed directly in an LB film on thiolated gold was inactive with respect to imide. The surface plasmon resonance measurements evidenced the stepwise assembly of complementary nucleotides on SAM/LB templates through consecutive phosphate-Zn2+-cyclen coordination. Base pairing between nucleotides resulted in a formation of A-U bilayers comprising two complementary monolayers. Finally, we report on SAM/LB patterns designed for divalent molecular recognition of uridine phosphate by amphiphilic bis(Zn2+-cyclen).     

On the origin of the unique properties of supported Au nanoparticles

The unique catalytic activity of supported Au nanoparticles has been ascribed to various effects including thickness/shape, the metal oxidation state, and support effects. Previously, we reported the synthesis of ordered Au monolayers and bilayers on TiO(x), with the latter being significantly more active for CO oxidation than the former. In the present study, the electronic and chemical properties of ordered monolayer and bilayer Au films have been characterized by infrared reflection adsorption spectroscopy using CO as a probe and ultraviolet photoemission spectroscopy. The Au overlayers are found to be electron-rich and to have significantly different electronic properties compared with bulk Au. The common structural features of ordered Au bilayers and Au bilayer nanoparticles on TiO2(110) are described, and the exceptionally high catalytic activity of the Au bilayer structure related to its unique electronic properties.     

Self‐assembly and photovoltaic properties of multilayer films based on partially doped polyaniline and poly(4‐carboxyphenyl)acetylene

Multilayer self‐assembled films, consisting of partially doped polyaniline (PAN) as a polycation and water‐soluble poly(4‐carboxyphenyl)acetylene (PCPA) as a polyanion, were fabricated through electrostatic attraction. These ultrathin PCPA/PAN films were relatively stable toward aqueous electrolyte solutions and polar organic solvents, and the photoelectric conversion properties of the self‐assembled PCPA/PAN films could be measured with traditional three‐electrode cells in 0.5 M KCl aqueous solutions. With an increase in the bilayer number, the photocurrent rose, and it reached its maximum at eight bilayers. A further increase led to a current drop due to an increase in the recombination probability and weak visible‐light transmission. When neutral PAN films were used, the photocurrent increased consecutively within 15 bilayers, and this indicated that the PAN molecules in the neutral state were stronger electron donors than those in the partially doped state. © 2004 Wiley Periodicals, Inc. J Polym Sci Part A: Polym Chem 42: 3224–3229, 2004     

Electrode nanomaterials self-assembled from thiolate-encapsulated gold nanocrystals

This paper describes the preliminary findings of an investigation of thin film assembly from monolayer-encapsulated gold nanocrystals and 1,9-nonanedithols. The creation of novel electrode nanomaterials derived from intriguing combinations of the encapsulating shells and the particle cores constitutes the motivation of this work. Narrow-sized, shaped and encapsulated nanocrystals were assembled as thin films on different substrates via an exchange reaction between alkanethiolates on the nanocrystal shells and dithiols in the solution. Both microscopic and spectroscopic data have confirmed the formation of dithiol-linked nanocrystals in the thin films. The electrochemical study has revealed interesting parallels and differences between monolayers on planar and nanocrystal gold surfaces, which have important implications to the correlation between binding properties at nanocrystal facets and the electrode properties of this interesting class of composite nanomaterials.     

Minimizing defects in polymer-based Langmuir-Blodgett monolayers and bilayers via gluing

Polymeric surfactants were prepared by quaternization of poly(4-chloromethylstyrene) with N,N-dimethyl-N-n-dodecylamine and N,N-dimethyl-N-n-octylamine to give 1 and 2, respectively. Each of these polymers formed stable monolayers at the air/water interface. Injection of poly(acrylic acid) (PAA) beneath the surface of these films led to a substantial increase in their cohesiveness (i.e., "gluing"), as evidenced by a dramatic increase in their surface viscosity. Examination of monolayers of 1 by atomic force microscopy, after being transferred to silicon wafers that were surface-modified with n-octadecyltrichlorosilane, showed that the presence of PAA leads to intact film. In contrast, transfer of unglued monolayers resulted in poor coverage. Comparison of the barrier properties of single glued and unglued LB bilayers formed in the presence and in the absence of PAA have shown that PAA minimizes defect formation within these ultrathin assemblies.     

Adsorption of hydrophobin proteins at hydrophobic and hydrophilic interfaces

The surface activity of two hydrophobin proteins, HFBII and SC3, at the solid–liquid, liquid–liquid and liquid–vapor interface has been investigated. Hydrophobins are fungal proteins that are known to adsorb and affect the physico-chemical properties of an interface. In this study, the surface activity was determined by measuring the interaction of hydrophobin molecules with various liquids, solid particles and films that are commonly used or produced in industrial processes. We found that a very low concentration of hydrophobin is required to facilitate the wet-in of hydrophobic solid particles, such as Teflon^®, into aqueous solutions. It is also capable of stabilizing aqueous dispersions of Kevlar^® nanopulp, reversing the wettability of hydrophobic films and stabilizing polyunsaturated fatty acid (PUFA) oil-in-water emulsions.     

Aggregation of a peptide antibiotic alamethicin at the air-water interface and its influence on the viscoelasticity of phospholipid monolayers

The aggregation properties of an antibiotic membrane-active peptide alamethicin at the air-water interface have been studied using interfacial rheology and fluorescence microscopy techniques. Fluorescence microscopy of alamethicin monolayers revealed a coexistence of liquid expanded (LE) and solid phases at the surface concentrations studied. Interfacial oscillatory shear measurements on alamethicin monolayers indicate that its viscoelastic properties are determined by the area fraction of the solid domains. The role of zwitterionic phospholipids dioleoylphosphatidyl choline (DOPC) and dioleoylphosphatidyl ethanolamine (DOPE) on the peptide aggregation behavior was also investigated. Fluorescence microscopy of alamethicin/phospholipid monolayers revealed an intermediate phase (I) in addition to the solid and LE phase. In mixed monolayers of phospholipid (L)/alamethicin (P), with increase in L/P, the monolayer transforms from a viscoelastic to a viscous fluid with the increase in area fraction of the intermediate phase. Further, a homogeneous mixing of alamethicin/lipid molecules is observed at L/P > 4. Our studies also confirm that the viscoelasticity of alamethicin/phospholipid monolayers is closely related to the alamethicin/phospholipid interactions at the air-water interface.     

Negative dipole potentials of uncharged langmuir monolayers due to fluorination of the hydrophilic heads

The dipole potential, affecting the structure, functions, and interactions of biomembranes, lipid bilayers, and Langmuir monolayers, is positive toward the hydrocarbon moieties. We show that uncharged Langmuir monolayers of docosyl trifluoroethyl ether (DFEE) exhibit large negative dipole potentials, while the nonfluorinated docosyl ethyl ether (DEE) forms films with positive dipole potentials. Comparison of the Delta V values for these ethers with those of the previously studied(37-39) monolayers of trifluoroethyl ester (TFEB) and ethyl ester of behenic acid (EB) shows that the reversal of the sign of Delta V causes the same change Delta(Delta V) = -706 +/- 16 mV due to fluorination of heads. The Delta V values of both TFEB and EB films differ by -122 +/- 16 mV from those of DFEE and DEE monolayers, respectively, with the same density. Such quantitative coincidence points to a common mechanism of reversal of the sign of the dipole potential for the ether and ester films despite the different structure of their heads. The mechanical properties and phase behaviors of these monolayers show that both fluorinated heads are less hydrated, suggesting that the change of the sign of Delta V could, at least partially, be related to different hydration water structure. The same negative contribution of the carbonyl bond in both TFEB and EB films contrasts with the generally accepted positive contribution of the C(delta+)=O(delta-) bond in condensed Langmuir monolayers of fatty acids, their alcohol esters, glycerides, and phospholipids but concurs with the theoretical analysis of Delta V of stearic acid monolayers. Both results question the literature values of the molecular dipole moments of these substances calculated via summation of bonds and atomic group contributions. Mixed monolayers of DFEE and DEE show smooth monotonic variation of Delta V from +450 to -235 mV, indicating a way for adjustment of the sign and magnitude of the dipole potential at the membrane-water boundary and regulation of such membrane behaviors as binding and translocation rate of hydrophobic ions and ion-carriers, adsorption and penetration of amphiphilic peptides, polarization of hydration water, and short-range repulsion. The interaction of the hydrophobic ions tetraphenylboron TPhB- and tetraphenylphosphonium TPhP+ with DFEE and DEE monolayers qualitatively follows the theory of binding of such ions to lipid bilayers, but the shifts Delta(Delta V) from the values obtained on water are much smaller than those for DPPC monolayers. This difference seems to be due to the solid (polycrystalline) character of the DFEE and DEE films that hampers the penetration of TPhB- and TPhP+ in the monolayers and reduces the attractive interaction with the hydrophobic moiety. This conclusion orients the future synthesis of amphiphiles with fluorinated heads to those which could form liquid-expanded Langmuir monolayers.     

原油活性组分油水界面膜扩张粘弹性研究

研究了用超临界萃取分馏法（SFEF）从伊朗重质原油中分离的两个具有不同平均分子量的原油界面活性组分在正癸烷/水界面的扩张粘弹性行为以及温度对体系扩张粘弹性的影响.研究发现平均分子量大的样品能在油水界面形成更为牢固的界面膜.从扩张模量幅度对扩张频率的双对数曲线和扩张模量相角的频率依赖关系可以推断所有实验体系界面膜的主要的弛豫过程不是扩散弛豫,而可能主要是通过吸脱附势垒的弛豫过程.温度对两个样品的扩张粘弹性参数都有强烈的影响.升高温度可以降低膜的强度和粘度,并且改变相角的频率响应.