Junbai Li

Effect of protein penetration into phospholipid monolayers: morphology and structure.

Abstract Phase transition and phase properties of dipalmitoylphosphatidylcholine (DPPC) monolayers penetrated by bovine β-lactoglobulin dissolved in a buffered aqueous subphase are experimentally studied. The phase transition during the penetration dynamics is indicated by a break point in the Π(t) transients. The condensed phase domains formed during the β-lactoglobulin penetration are visualized by Brewster angle microscopy (BAM). The lattice structure of the condensed phase is characterised by grazing incidence X-ray diffraction (GIXD). Experiments on the penetration kinetics of β-lactoglobulin into DPPC monolayers are performed, starting from different monolayer states and using different protein concentrations. The condensed phase formed after the main phase transition point, consists only of DPPC. The β-lactoglobulin penetration occurs without any specific interaction with the DPPC molecules. Number and growth of the domains depend on the area per DPPC molecule at which the β-lactoglobulin penetration takes place. A first-order main phase transition can be induced when the protein penetrates into a fluid (gaseous) DPPC monolayer. β-Lactoglobulin cannot penetrate into a condensed DPPC monolayer at a surface pressure above the equilibrium penetration pressure. Conformational changes and squeezing out of protein from the penetrated monolayer are studied by compression of penetrated monolayers in equilibrium.

The aggregation and phase separation behavior of a hydrophobically modified poly(N-isopropylacrylamide).

Abstract To study the water solution properties of a hydrophobically modified poly( N -isopropylacrylamide) (PNIPAM) that is temperature-sensitive, the copolymer of N -isopropylacrylamide (NIPAM) and octadecylacrylate (ODA) was synthesized. The aggregation behavior of the copolymer was studied by surface tension and fluorescent probe methods. Simultaneously, the phenomenon of LCST (Lower Critical Solution Temperature) of the copolymer in aqueous solution with the increase of temperature was also studied by using the fluorescent probe method. Results showed that phase separation occurred in aqueous solution of the copolymer when the temperature was increased to its LCST. The π – A isotherms for the copolymer molecules, as an insoluble monolayer on the water–air interface, was determined by the Langmuir–Blodgett method. The abnormal phenomenon, which the monolayer of the copolymer molecules became more condensed and more condensed with the increase of temperature, was observed. It further indicated that phase separation of the copolymer occurred from another angle. In addition, to prove the thermo-sensitive effect of the copolymer on the release behavior of liposomes, small unilamellar vesicles entrapped with 5(6)-carboxyfluorescein (5(6)-CF) were coated with the copolymer. We found that the coating of the copolymer resulted in reduction of the release below 30°C and enhancement of the release above 30°C. It indicates that there are obvious interactions between the copolymer and liposomes.

Stabilized complex film formed by co-adsorption of β-lactoglobulin and phospholipids at liquid/liquid interface

Abstract A complex film has been formed at a drop surface by the co-adsorption of β-lactoglobulin with the l -α-dipalmitoylphosphatidylcholine (DPPC), l -α-dipalmitoylphosphatidyl-ethanolamine (DPPE), and l -α-dipalmitoylphosphatidic acid sodium salt (DPPA), respectively, at the water/chloroform interface. By using pendent drop technique we studied the headgroup effect of lipids on the kinetics of co-adsorption layers. It was found experimentally that a folded drop surface was formed during co-adsorption and the headgroup of lipids affected on the formation rate of the folded drop surface. Such a skinlike film demonstrates that there exits an strong interaction between β-lactoglobulin and each lipid. By depositing one lipid mono- and bilayers onto a solid surface morphology of the mixed lipid/β-lactoglobulin layers has been investigated by atomic force microscopy (AFM).

Dynamic characterization of phospholipid/protein competitive adsorption at the aqueous solution/chloroform interface.

Abstract In the present paper, we use the drop volume method to study the dynamics of the competitive adsorption of the zwitterionic phospholipids (DPPC, DPPE, DMPC and DMPE) mixed with proteins (β-lactoglobulin, β-casein, and human serum albumin, respectively) at the chloroform/water interface. In order to investigate the main factors influencing the equilibrium interfacial tension of the mixed system (drops of lipid in chloroform formed in an aqueous protein solution environment), proteins of different conformation and concentration, and phospholipids of different structure have been investigated. It is observed that, with constant external protein concentration, the equilibrium interfacial tension γ decreases with the increase of internal lipid concentration. When the phospholipid concentration is close to the CAC, both the conformation and concentration of the protein do not influence the equilibrium interfacial tension of the mixed systems remarkably. With the same internal phase containing phospholipid in oil solvent and different external phases containing the protein in water, the γ– C isotherms show similar tendencies. Moreover, the structure of the phospholipid determines the equilibrium interfacial tension, where the lipid head group is much more significant rather than the chain length. The experimental results show that in DMPC-protein systems, the equilibrium interface tension decreases with the phospholipid concentration more rapidly than in DMPE-protein systems.

Stability investigation of the mixed DPPC/protein monolayer at the air–water interface

Abstract A subphase exchange method has been used to investigate the stability of the mixed l -α-dipalmitoylphosphatidylcholine (DPPC)/β-lactoglobulin monolayer formed via the protein adsorption into the spread lipid layer at the air–water interface by Brewster angle microscopy (BAM). The BAM images demonstrated that the adsorbed protein might compress the DPPC monolayer to enter a phase transition early close to a condensed phase. When protein concentration is very high the penetration of the protein is rather fast, and a larger domain was preferentially formed without compressing the DPPC monolayer. After the mixed DPPC/β-lactoglobulin monolayer was formed, the protein solution in subphase was totally washed out by pumping in buffer water for a long time. In the fluid phase and coexistence region of DPPC monolayer with penetrated protein layer the domains have been recorded by BAM images for comparing the variation before and after the exchange of subphase. The experimental results revealed that there was not significant change of the domains in size and shape. It indicates that the adsorbed protein has a strong interaction with DPPC and is more likely to remain at the interface. A relatively stable mixed lipid–protein monolayer can be constructed by this way. Hydrophobic and electrostatic interactions between DPPC and β-lactoglobulin as well as the conformation change of β-lactoglobulin at air–water interface have been taken into account to stabilize the mixed layers.

Structure characterization and stability of mixed lipid/protein monolayer at the air/water interface.

Abstract A mixed protein/lipid monolayer has been constructed by the protein adsorption from subphase into the spread phospholipid monolayer. A precisely controlled pump was used to exchange the protein solution with different pH values after the protein was ensured to reach the less condensed surface. The domains formed in the coexistence region of D-dipalmitoylphosphatidylcholine (D-DPPC) have been recorded by Brewster angle microscopy (BAM) combined with the film balance before and after the penetration of the protein, human serum albumin (HSA). The subphase was exchanged by gradually increasing or decreasing pH value of the solution. Three isotherms of the mixed D-DPPC/HSA monolayer with the subphase of pH=4.2, pH=7.0 and pH=9.1, respectively, were obtained. It indicated that the area per lipid molecule with protein increased as the subphase pH value was lowed. Simultaneously, morphological dynamic changes caused by the gradual changing on subphase pH were observed. These variations can be ascribed to the conformation change of protein under the fluctuation of pH value. The hydrophobic and electrostatic interactions between the phospholipid and HSA were as considered for the interpretation of domain change based on the current experimental results

pH value and ionic strength effects on the adsorption kinetics of protein/phospholipid at the chloroform/water interface.

The adsorption kinetics of β-lactoglobulin in an aqueous buffer solution in the presence of phospholipid, l-α-dipalmitoylphosphatidylethanolamine (DPPE), dissolved in chloroform, has been investigated by using a pendent drop technique. The dynamic interfacial tension for the mixed β-lactoglobulin/phospholipid layer was measured as a function of time at a constant phospholipid concentration, CDPPE=1×10−6 M. Experimentally it has been found that increase of ionic strength of aqueous solution can enhance the adsorption rate of the protein. However, the larger pH value leads to the adsorption process being reduced. When the protein concentration was increased up to 3.8 mg l−1, the effect of the salt concentration on the adsorption can be neglected and the protein dominates principally the adsorption process. In the acid aqueous solution, pH 5, approaching equilibrium time of adsorption for protein was around 1300 s and twice longer than in the base subphase, pH 8. However, the equilibrium interfacial tension was independent on the pH value. The salt concentration and pH value effects show that both the electrostatic interaction and hydrophobic effect make the mixed lipid/β-lactoglobulin layer stable at the liquid/liquid interface.

Effect of dodecyl dimethyl phosphine oxide penetration into phospholipid monolayers: morphology and dynamics

Abstract The penetration kinetics of dodecyl dimethyl phosphine oxide (C 12 DMPO) into a Langmuir monolayer of dipalmitoylphosphatidylcholine (DPPC) and the corresponding morphology were experimentally studied. It was found that under proper conditions the penetration of dodecyl dimethyl phosphine oxide could induce a first-order phase transition in the fluid-like lipid Langmuir monolayer. The transition is indicated by a break point in the Π ( t ) penetration kinetics curves and can be visualized by BAM. The dynamics of the penetrated layer and the subphase effect on the Langmuir monolayer were also investigated. The experimental results show that the adsorption of soluble molecules increases the surface density, which in turn causes the first-order phase transition in the monolayer. When the surface density reaches a certain critical value, the main phase transition sets in.