Surface-active Proteins Research Articles

Surface-active proteins enable microbial aerial hyphae to grow into the air.

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Structure and function of surfactant protein B and C in lipid monolayers: a scanning force microscopy study

This study focuses on the impact of surfactant protein C and B on lipid monolayers at various surface pressures. The artificial system is composed of the saturated phospholipids dipalmitoylphosphatidylcholine (DPPC) and dipalmitoylphosphatidylglycerol (DPPG) in a molar ratio of 4:1 with 0.2 mol% SP-B and/or 0.4 mol% SP-C. A dominating influence of SP-C on the morphology of lipid monolayers at high surface pressure was found. Even in the presence of both proteins structural peculiarities typical for SP-C were found at elevated pressure employing tapping mode scanning force microscopy of LB-films. Stacked bilayer-protrusions known to be induced by SP-C are visible in films containing SP-C together with SP-B. The findings were corroborated by fluorescence microscopy at the air/water interface and are consistent with the appearance of the corresponding isotherms. In the low pressure regime, however, disc-like protrusions characteristic of SP-B containing films are discernible. Filamentous LE domains with large boundaries arise due to the reduced line tension in the presence of surface active proteins, particularly SP-B. Remarkably, SP-B fluidizes the monolayer to a larger extent than SP-C as revealed by scanning force microscopy images. These findings show that SP-B and SP-C interact independently of each other. Therefore we conclude that SP-C may be responsible for the fast respreading process during the breathing cycle while SP-B removes material from the monolayer in more discrete portions.

Effects of lecithins and proteins on the stability of emulsions

Food consists of three components, carbohydrates, proteins and fats, which are linked by various types of chemical and physical bonds. The interactions at the interface of dispersions and emulsions are influenced by surface-active emulsifiers such as lecithins. Synergistic effects on the emulsion stability can be obtained by selected protein-lecithin combinations. Hydrocolloids enhance the stability by viscosity increase and gel formation. Lecithins are modified physically and enzymatically, giving a range of food grade emulsifiers with different hydrophilic-lipophilic-balance (HLB) values. The influence of phospholipid fractions in the homogenization process can be measured by the particle size distribution technique (PSD) and emulsifying tests, which assess the emulsion stability. Basic studies on the interactions of surface-active proteins and lecithins in emulsions are reviewed. The principles of combined use of proteins and lecithins are presented for processed foods such as mayonnaise, margarine. instant milk powders. fat reduced cookies, and chocolate coatings for ice cream dipping. A new challenge for the food industry is the liposome encapsulation technique. Liposomes are produced with phosphatidylcholine fractions to encapsulate water and oil soluble components, such as flavours, in one capsule.

Prediction of separation factor in foam separation of proteins

A phenomenological model has been developed for predicting separation factors obtained in concentrating protein solutions using batch-foam columns. The model considers the adsorption of surface active proteins onto the air-water interface of bubbles, and drainage of liquid from the foam, which are the two predominant processes responsible for separation in foam columns. The model has been verified with data collected on casein and bovine serum albumin (BSA) solutions, for which adsorption isotherms are available in the literature. It has been found that an increase in liquid pool height above the gas distributor and the time allowed for drainage result in a better separation. Further, taller foam columns yield poorer separation at constant time of drainage. The model successfully predicts the observed results.

Dynamic Penetration of an Insoluble Monolayer by a Soluble Surfactant: Theory and Experiment

Surfactants or surface active proteins adsorbing onto aqueous−gas interfaces can require many minutes to hours to attain equilibrium, depending upon the size, bulk concentration, and chemical structure of the adsorbing species. In contrast, soluble surface active molecules adsorbing into an insoluble monolayer equilibrate on a time scale that is more rapid. Here, the time dependent behavior of the adsorption of a soluble surface active molecule into an insoluble monolayer is addressed using a diffusion-controlled adsorption model with a Langmuir or Frumkin adsorption isotherm. In the Langmuir framework, the presence of an insoluble monolayer occupying the fraction x1 of the interface reduces the amount of soluble surfactant adsorbed at equilibrium by the factor (1 − x1). Because less surfactant must be delivered to the interface, the equilibration time scale for the adsorption process is reduced by the factor (1 − x1)2 as compared to adsorption onto a clean interface. In a Frumkin framework, the role of c...

The Effect of Surface Active Additives on the Partitioning of Proteins and Enzymes in Aqueous Two‐Phase Systems

The effect of cationic and anionic surface active additives on the partitioning of proteins has been studied. The surface active agents interact with proteins electrostatically and also hydrophobically. The increased hydrophobicity of the proteins because of association with surfactants is responsible for the changes in the partition coefficients and in some cases for precipitation of the proteins. Specific interactions also exist between mildly surface active amino acids and proteins.

The Effect of Surface Active Additives on the Partitioning of Proteins and Enzymes in Aqueous Two-Phase Systems

The effect of cationic and anionic surface active additives on the partitioning of proteins has been studied. The surface active agents interact with proteins electrostatically and also hydrophobically. The increased hydrophobicity of the proteins because of association with surfactants is responsible for the changes in the partition coefficients and in some cases for precipitation of the proteins. Specific interactions also exist between mildly surface active amino acids and proteins.

Purification and characterization of a 28 kDa cytosolic inhibitor of cholesteryl ester hydrolases in rat testis.

A 28 kDa inhibitory protein was purified from rat testis cytosol by sequential 40-65% ammonium sulfate precipitation, cation exchange chromatography, anion exchange chromatography, and preparative SDS-polyacrylamide gel electrophoresis. The heat-stable, trypsin-labile protein exhibited nonenzymatic, concentration-dependent inhibition of testicular and pancreatic cholesteryl ester hydrolases at all stages of purification. Copurifying at each stage was a 26.5 kDa protein which comprised 25% of the mass of the two proteins. Polyclonal antibodies raised to either or both 28 kDa and 26.5 kDa proteins by direct injection of excised electrophoretic bands cross-reacted with both proteins on western blots, immunoprecipitated both proteins, and neutralized inhibitory activity. Amino acid compositions of the individual proteins electroeluted from SDS-polyacrylamide gels were different from those of other surface-active proteins of similar molecular weights. Both proteins exhibited identical pl of 4.8 on chromatofocusing columns and two-dimensional gel electrophoresis. Although the subcellular distribution of the 28 kDa protein is unknown, its testicular cytosolic concentration, calculated from the purified protein mass, was 8 X 10(-9) mols/L, which probably underestimates the actual concentration by an order of magnitude. This is greater than the minimum concentration required for in vitro inhibition (10(-9) mols/L), consistent with a physiological role for this protein.

Behavior of insect defensin A at the air/water interface

The surface behavior of insect defensin A, an inducible antibacterial protein isolated from the larvae of the fleshfly Phormia terranovae, has been investigated. Defensin A is able to adsorb at the air/water interface from an aqueous solution. The adsorption kinetics show two regions corresponding to adsorption and interfacial rearrangements. Both the maximum pressure reached in the course of adsorption and the equilibrium spreading pressure, π e, rank defensin A among surface-active proteins. The data extracted from the pressure-area (π-A) isotherms suggest that the defensin A molecule is partly unfolded at the air/water interface and may self-associate under compression. The stability of the defensin monolayer is greatest at basic pH (around the isoelectric point) and increases with increase in the ionic strength of the subphase. The presence of 2 mM Ca 2+ ions, pH 7.2, gives rise to an increase in the monolayer compressibility. Compression-expansion cycles present a wide hysteresis interpreted as the result of simultaneous desorption, self-association and folding of the defensin molecules in the course of the compression process.

Biosensors: Adaptation for practical use

This paper explores the interfacing of biosensors with biological fluids. The need for performance to be independent of solution variables is first emphasized, so that devices can be used without a sample preparation step. The behaviour of surface-active proteins and cells is then summarized, and how these can lead to altered biosensor responsiveness. Strategies for adapting biosensors for long-term use within biological matrices are suggested.

A comparison of the surface activities of human apolipoproteins A-I and A-II at the air/water interface

Surface pressure (π) and adsorption isotherms for human apolipoproteins A-I and A-II at the air/water interface have been determined and used to deduce the probable molecular structures of the monomolecular films. The surface concentrations were measured using the surface radioactivity method to monitor the adsorption of reductively [ 14C]methylated apoproteins. Apolipoprotein A-I and apolipoprotein A-II are extremely surface-active proteins and adsorb to exert maximal π values of 22 and 24 mN · m −1 respectively, at a steady-state subphase concentration of about 3 · 10 −5 g/100 ml (equivalent to 11 and 17 nM for apolipoprotein A-I and apolipoprotein A-II, respectively). At saturation monolayer coverage, the average molecular areas for apolipoprotein A-I and apolipoprotein A-II are 15 and 13 Å 2/residue, respectively. These packing densities are consistent with monolayers consisting largely of α-helical protein molecules lying with the long axes of the helical segments in the plane of the interface. Comparison of the molecular packings of spread and adsorbed monolayers of these proteins indicates that at low π values, the adsorbed films are more expanded, but at high π values, the molecular packing in both types of film is the same.

Functional Characteristics of the Mucilaginous Polysaccharides Derived from Cowpea (Vigna sinensis), Black Gram (Phaseolus mungo) and Linseed (Linum usitatissimum)

AbstractThe cowpea polysaccharides had the maximum viscosity at around pH 6 and it decreased with increase in temperature but increased with increase in concentration. Both black gram and cowpea polysaccharides stabilized against thermal disruption the foam formed by the surface active proteins. Black gram starch and the mucilage together showed greater foam stability, probably due to enhanced viscosity caused by synergistic interactions between these polysaccharides. Gelatinization temperature as well as the peak and set back viscosities of wheat starch were considerably decreased by adding either the black gram or the linseed polysaccharide. In farinograph experiments the black gram polysaccharide exhibited a strengthening effect on wheat flour dough, whereas the linseed polysaccharide resulted only in an increased water absorption. Extensographic data indicated increased resistance and decreased extensibility of wheat flour dough by the addition of black gram and linseed polysaccharides.

Design and characterization of peptides with amphiphilic beta-strand structures.

To extend our studies on peptides and proteins with amphiphilic secondary structures, a series of peptides designed to form amphiphilic beta-strand structures was designed, synthesized, and characterized by circular dichroism and infrared spectroscopy. Amphiphilic beta-strand conformations may be likely to appear in a variety of surface-active proteins, including apolipoprotein B and fibronectin. In a beta-strand conformation, the synthetic peptides will possess a hydrophobic face composed of valine side chains and a hydrophilic face composed of alternating acidic (glutamic acid) and basic (ornithine or lysine) residues. The peptides studied had a variety of chain lengths (5, 9, and 13 residues), and had the amino groups either free or protected with the trifluoroacetyl group. While the peptides did not possess a high potential for beta-sheet formation based on the Chou Fasman parameters, they possessed significant beta-sheet content, with up to 90% beta-sheet calculated for the 13-residue protected peptide. The driving force for beta-sheet formation is the potential amphiphilicity of this conformation. The beta-strand conformation of the 13-residue deprotected peptide was stable in 50% trifluoroethanol, 6 M guanidine hydrochloride, and octanol. The peptides are strongly self-associating in water, which would reduce the unfavorable contacts of the hydrophobic residues with water. It is clear that small peptides can be designed to form stable beta-strand conformations.

Phase behavior of enzymatically modified proteins in concentrated aqueous systems.

Enzymatically modified proteins produced from gelatin by papain-catalyzed incorporation of leucine hexyl ester and leucine dodecyl ester, designated as EMG-6 and EMG-12, respectively, are known to be potent surfactants [Agric. Biol. Chem., 46, 173 (1982)]. In the present study, experiments were carried out to characterize the phase behavior of these surface-active proteins in concentrated aqueous systems at elevated temperatures. For 60%- and 70%-EMG products, it was found that their elasticity and viscosity as represented by the storage modulus (G') and loss modulus (G), respectively, decreased greatly at 30°C and 80°C. Polarized microscopic observations showed that in the temperature range of 30-80°C, a liquid crystalline structure of a lamel ar type was formed. Differential thermal analysis demonstrated that, in accordance with the liquid crystal formation, an endothermic process took place, which gives evidence for an energetic change associated with this phase transition. Such phase transition phenomena were not observed for a gelatin hydrolysate, bovine serum albumin and casein used as controls.

Kinetic model for surface-active enzymes based on the Langmuir adsorption isotherm: phospholipase C (Bacillus cereus) activity toward dimyristoyl phosphatidylcholine/detergent micelles.

A simple kinetic model for the enzymatic activity of surface-active proteins against mixed micelles has been developed. This model uses the Langmuir adsorption isotherm, the classic equation for the binding of gas molecules to metal surfaces, to characterize enzyme adsorption to micelles. The number of available enzyme binding sites is equated with the number of substrate and inhibitor molecules attached to micelles; enzyme molecules are attracted to the micelle due to the affinity of the enzyme active site for the molecules in the micelle. Phospholipase C (Bacillus cereus) kinetics in a wide variety of dimyristoyl phosphatidylcholine/detergent micelles are readily explained by this model and the assumption of competitive binding of the detergent at the enzyme active site. Binding of phospholipase C to pure detergent micelles is demonstrated by gel filtration chromatography. The experimentally determined enzyme-detergent micelle binding constants are used directly in the rate equation. The Langmuir adsorption model predicts a variety of the characteristics observed for phospholipase kinetics, such as differential inhibition by various charged, uncharged, and zwitterionic detergents and surface-dilution inhibition. The essential idea of this model, that proteins can be attracted and bound to bilayers or micelles by possessing a binding site for the molecules composing the surface, may have wider application in the study of water-soluble (extrinsic) protein-membrane interactions.

Anticomplementary Activity of Human Immunoglobulin G.

The anticomplementary activity of IgG can be increased up to 20-fold by pipetting during the preparation of serial dilutions for the assay of this activity. Albumin, if added to the IgG solution before the serial dilutions, completely prevents this artifactual increase in activity. Polyethylene glycol, polyvinyl pyrrolidone, methyl cellulose, gelatin and octanol are also effective stabilizers of IgG. Repeated pipetting of IgG solutions caused marked linear increase in their anticomplementary activity. The formed anticomplementary activity was due to small amounts of highly aggregated protein. The amount of activity formed depended on at least four factors: [1] the number of pipetting steps; [2] the IgG concentration; [3] the level of albumin or other stabilizers, and [4] the pH, which influences the stabilization by albumin. The anticomplementary activity of IgG is increased up to 100-fold by exposure to gas-liquid, liquid-liquid and hydrophobic solid-liquid interfaces. Albumin and polyethylene glycol prevent the activity increase during these treatments. The tendency of IgG to aggregate at interfaces and the ability of albumin and other substances to prevent the aggregation paralleled their rate of adsorption to the air-water interface. Solutions of dansyl chloride in decane emulsified in aqueous solutions of IgG and albumin specifically label the proteins at the liquid-liquid interfaces. The mechanism of stabilization can be explained by preferential adsorption of surface-active proteins and polymers.

Interaction between Surface Active Agents and Proteins. IV. Heat of Interaction of Sodium Dodecyl Sulfate and Egg Albumin

Abstract We measured the heat evolved when egg albumin and SDS solutions were mixed at pH 6.8 and at 25.00°C, and calculated the heat of interaction.

Interaction between Surface Active Agents and Proteins. III. Precipitation Curve of the System Sodium Dodecyl Sulfate-Egg Albumin at Various pH’s and the Determination of the Concentration of Protein by the Titration Using Surfactant

Abstract (1) The precipitation curve of the system egg albumin-SDS was studied in the pH region between 4.2 and 2.3, and at constant ionic strength 0.30. It was found that the number of SDS molecules bound to one molecule of egg albumin is constant and this is considered to be a verification of the fact that the number of positive charges on one egg albumin molecule is constant in this pH region. (2) A rapid determination of the protein concentration by the titration using SDS is possible within an error of ±2%. (3) Egg albumin and horse serum albumin react in different ways with SDS in distilled water and in sodium chloride solution.

Interaction between Surface Active Agents and Proteins. II. Electrophoretic Investigation of the System Sodium Dodecyl Sulfate and Egg Albumin

Abstract We studied electrophoretically the interaction of SDS and egg albumin in the pH range between 5.4 and 10.8, higher pH’s than that of the isoelectric point of egg albumin. Results obtained are as follows. (1) The electrophoretic patterns of the system egg albumin-SDS can be divided into three groups. This method of division is different from that in the system SDS-horse serum albumin by Putnam and Neurath. The first group of the division contains the patterns obtained when the mixing ratio albumin/SDS is between 100/0 and 80/20, and here the complex ADn (n=40, A: albumin, D : detergent) and egg albumin coexist. The second group contains the patterns obtained when the mixing ratio albumin/SDS is between 80/20 and 35/65 and in this region the composition of the complex changes continuously from ADn to AD8n, The third region appears when the mixing ratio albumin/SDS is between 35/65 and 0/100 and here the complex ADSn and SDS coexist. The above mixing ratio values are obtained at pH 6.8, and they change somewhat as a function of pH. (2) Below pH 7, a number n of the first complex ADn equals the number of positive charges on one egg albumin, but above pH 7, this relation cannot hold. (3) In the reaction between egg albumin and SDS, it is not assumed that SDS reacts only in a micellar form but that SDS existing as a single ion can react.

Interaction between Surface Active Agents and Proteins. I. Precipitations formed by Mixing Sodium Alkyl Sulfates and Egg Albumin

Abstract (1) Precipitation reactions between SDS and egg albumin, SOS and egg albumin, and SBS and egg albumin were studied. There was an instant precipitation when SDS and egg albumin solutions were mixed, and when SOS and egg albumin solutions were mixed, but not when SBS and egg albumin solutions were mixed. Comparing the precipitation curve of the SDS-egg albumin system and that of the SOS-egg albumin system, we can find that both detergents form a complex having the same formula AD40 (A : albumin, D: detergent) in the region of protein excess, and therefore we can conclude that there is no difference between the reactivities of SDS and SOS toward egg albumin. On the contrary, in the region where redissolution of precipitate occurs, the precipitation curves do not coincide. This means that the two detergents have different powers to dissolve precipitate. In addition, when the concentration of SOS differs, the power to dissolve precipitate differs. (2) It was found electrophoretically that the complex is formed in turbid supernatants after centrifugation and also in transparent solutions the composition of which is in the region where no precipitation occurs. (3) Observing the precipitation curves of the system horse serum albumin-SDS and the system egg albumin-SDS, we can see that there is a region where no precipitation occurs when a great excess of serum albumin is used; but there is not such a region when a great excess of egg albumin is used. (4) We measured the lowering of the CMC of SDS and of SOS caused by the increase in the concentration of buffer solution. The lowering does not depend upon the kinds and pH’s of buffer solutions but upon the ionic strength of buffer solution which dissolves detergent.