Micellization of Surfactin and Its Effect on the Aggregate Conformation

The results show that surfactin has strong self-assembly ability to form micelles and the micelles tend to form larger aggregates. Surfactin adopts a ...
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J. Phys. Chem. B 2008, 112, 15195–15201

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Micellization of Surfactin and Its Effect on the Aggregate Conformation of Amyloid β(1-40) Yuchun Han, Xu Huang, Meiwen Cao, and Yilin Wang* Key Laboratory of Colloid and Interface Science, Institute of Chemistry, Chinese Academy of Sciences, Beijing 100190, People’s Republic of China ReceiVed: July 7, 2008; ReVised Manuscript ReceiVed: August 25, 2008

The aggregation of amyloid β-peptide (Aβ(1-40)) into fibrils is a key pathological process associated with Alzheimer’s disease. This work has investigated the micellization process of biosurfactant surfactin and its effect on the aggregation behavior of Aβ(1-40). The results show that surfactin has strong self-assembly ability to form micelles and the micelles tend to form larger aggregates. Surfactin adopts a β-turn conformation at low micelle concentration but a β-sheet conformation at high micelle concentration. The effect of surfactin on the Aβ(1-40) aggregation behavior exhibits a strong concentration-dependent fashion. Below the critical micelle concentration of surfactin, the electrostatic binding of surfactin monomers on Aβ(1-40) causes Aβ(140) molecules to unfold. Assisted by the hydrophobic interaction among surfactin monomers on the Aβ(140) chain, the conformation of Aβ(1-40) transfers to the β-sheet structure, which promotes the formation of fibrils. At low surfactin micelle concentration, besides the electrostatic force and hydrophobic interaction, hydrogen bonds formed between surfactin micelles and adjacent Aβ(1-40) peptide chains may promote the ordered organization of these Aβ(1-40) peptide chains, thus leading to the formation of β-sheets and fibrils to a great extent. At high surfactin micelle concentration, the separating of Aβ(1-40) chains by the excessive surfactin micelles and the aggregation of the complexes of Aβ(1-40) with surfactin micelles inhibit the formation of β-sheets and fibrils. Introduction Surfactin, a kind of acidic lipopeptide excreted by various strains of Bacillus subtilis, is an attractive and powerful biosurfactant because of its special amphiphilic character.1 It consists of a peptide loop of seven amino acid residues (GluLeu-D-Leu-Val-Asp-D-Leu-Leu) and a C13-15 β-hydroxy hydrophobic fatty acid chain (Figure 1). The peptide ring of surfactin shows a “horse-saddle” topology in aqueous solution. The two negatively charged amino acid residues Glu and Asp constitute a minor polar domain. On the opposite side, Val residue extends down, facing the fatty acid chain, making up a major hydrophobic domain.2 Due to the special amphiphilic character, surfactin can remarkably lower the surface tension of water from 72 to 27 mN · m-1 at a concentration as low as 20 µM.1 As a biosurfactant, surfactin exhibits many interesting bioactive properties, including antiviral,3 antitumor,4 antibiotic,5 antibacterial,6 and hemolytic activities.7 In addition, due to its less toxic and better biodegradable properties than synthetic compounds, surfactin is receiving attention for industrial, biotechnological, and therapeutical applications. Alzheimer’s disease (AD) is a progressive neurodegenerative disorder characterized by a large number of amyloid deposits in the form of senile plaques in the brain.8 These plaques are primarily formed by amyloid β-protein (Aβ) of 39-42 amino acid residues, a proteolytic fragment of the larger amyloid β-protein precursor (APP).9 It is commonly considered that the fibrillar Aβ aggregate is an important reason in the etiology of AD, whereas recent investigations show that soluble Aβ aggregates (also called oligomers or protofibrils) may also be responsible for synaptic dysfunction in AD.10,11 Therefore, * To whom correspondence should be addressed. E-mail: yilinwang@ iccas.ac.cn.

Figure 1. The chemical structure of surfactin.

investigating the factors involved in Aβ fibrillogenesis is important to understand this neurodegenerative disease. The effect of surfactants on the fibrillation of Aβ(1-40) has been extensively studied during the past decades since surfactant micelles can mimic the biological membrane environment.12-17 Sabate´ and Estelrich13 reported the interaction of Aβ(1-40) with three alkylammonium bromides and found that the surfactants promoted Aβ(1-40) aggregation below the critical micelle concentration (CMC), whereas the presence of micelles induced a delay of the aggregation. Several authors have focused on the effect of SDS on the Aβ fibrillogenesis. Rangachari et al.17 found that the addition of SDS accelerated aggregation over a narrow range of SDS concentration. Jeng et al.18 found that SDS could suppress the fibril formation of Aβ(1-40) via the formation of small peptide/SDS complex in aqueous solutions. This suppression effect is enhanced by SDS micelles when a larger SDS concentration is used. In the previous works,19,20 we investigated the interaction of cationic gemini surfactant hexamethylene-1,6-bis(dodecyldimethylammonium bromide) (C12C6C12Br2) and Aβ(1-40) and found that C12C6C12Br2 had a strong ability to induce the fibrillar formation at very low surfactant concentrations because of its double charges and double hydrophobic chains. In this study, the interaction of surfactin with Aβ(1-40) has been studied. Compared with conventional synthetic surfactant, surfactin has a large ring with seven amino acid residues, which may contribute strong hydrogen bonds to the interaction between

10.1021/jp805966x CCC: $40.75  2008 American Chemical Society Published on Web 11/05/2008

15196 J. Phys. Chem. B, Vol. 112, No. 47, 2008 surfactin and Aβ(1-40). On the other hand, surfactin has a relatively large hydrophobic domain and carries two negative charges at pH 7.4 because the pKa values of Asp and Glu are around 4.3 and 4.5, respectively.21 Thus, both electrostatic force and hydrophobic interaction may also play important roles in the surfactin-Aβ(1-40) interaction. In addition, surfactin is usually considered as more effective, more environment friendly, and more stable than many synthetic compounds. These aspects inspire us to explore whether surfactin has a special effect on Aβ fibrillogenesis. In order to understand the interaction of surfactin with Aβ(1-40), the micellization behavior of surfactin has also been studied. The results indicate that surfactin of low concentration can induce a prominent change of Aβ(1-40) in its secondary structure and morphology; conversely, surfactin of high concentration cannot induce the secondary structure change and fibril formation of Aβ(1-40). Experimental Section Materials. Amyloid β(1-40) (trifluoroacetate salt) was purchased from Sigma. (Aβ(1-40), No. A4473, Lots 043K1348, 085K1804, and 095K1569). The purity was greater than 95%. The molecular weight of Aβ(1-40) was about 4328.9 g/mol. Surfactin (approximately 98% purity) was obtained from Sigma. 1,1,1,3,3,3-Hexafluoro-2-propanol (HFIP) was from ACROS. Chloroform and methanol were from Beijing Chemical Factory. All of the experiments were carried out in phosphate buffer of pH 7.4 with an ionic strength (I) of 10 mM. Pure water was obtained from Milli-Q equipment. Surfactin and Aβ(1-40) Sample Preparation. A stock solution of surfactin was prepared in chloroform/methanol (2: 1) and stored at -20 °C. Chloroform and methanol were removed by evaporation under a gentle stream of nitrogen just before surfactin was dissolved in phosphate buffer of pH 7.4. Aβ(1-40) in nonaggregated form was prepared as described previously.19 Briefly, a stock solution was prepared by dissolving 1.2 mg of Aβ(1-40) in 1.2 mL of HFIP, which was incubated at room temperature for 1 h in sealed vials. Then, the solution was bath-sonicated for 30 min until Aβ(1-40) dissolved completely. After that, the solution was kept at 4 °C for 30 min to avoid solvent evaporation when aliquoting. After aliquoted, the samples in HFIP were stored at -20 °C for later use. Just before initiating the following experiments, an aliquot of Aβ(1-40) was chosen, and HFIP was removed by evaporation under a gentle stream of nitrogen, leaving a slightly yellow film. Then, the sample was dissolved in phosphate buffer to reach a final Aβ(1-40) concentration of 40 µM. Isothermal Titration Microcalorimetry (ITC). The calorimetric measurements were conducted using a TAM 2277-201 microcalorimetric system (Thermometric AB, Ja¨rfa¨lla, Sweden) with a stainless steel sample cell of 1 mL. The cell was initially loaded with 0.6 mL of buffer or a 40 µM Aβ(1-40) buffer solution. The concentrated surfactin solution was injected into the sample cell via a 500 µL Hamilton syringe controlled by a 612 Thermometric Lund pump. A series of injections were made until the desired range of concentration had been covered. The system was stirred at 50 rpm with a gold propeller. The observed enthalpy (∆Hobs) was obtained by integration over the peak for each injection in the plot of heat flow P against time t. All of the measurements were conducted at 303.15 ( 0.01 K. Atom Force Microscopy (AFM). AFM morphology images were recorded using a tapping mode (Nanoscope IIIa multimode system, Digital Instruments, Santa Barbara, CA) with silicon cantilever probes in air. To prepare the specimen for AFM imaging, we incubated 3-5 µL aliquots of Aβ(1-40) solutions

Han et al. in the absence and presence of surfactin on the surface of freshly cleaved mica, usually for 10-15 min. Following incubation, the excess solution was briefly rinsed with distilled water and dried with a gentle stream of nitrogen. All of the measurements were conducted at room temperature. Analysis of the images was carried out using the Digital Instruments Nanoscope software (version 512r2). Circular Dichroism (CD). CD spectra were recorded on a JASCO J-815 in a 1 mm path length cell. Scans were obtained in a range between 190 and 260 nm by taking points at 0.5 nm, with an integration time of 0.5 s. Eight scans were averaged and smoothed to improve the signal-to-noise ratio. For surfactin, the resulting optical activities were reported as molar residue ellipticity [θ]MRW (deg · cm2 · dmol-1). For the mixtures of surfactin and Aβ(1-40), the resulting optical activities were reported as the ellipticity (mdeg). Transmission Electron Microscopy (TEM). Samples were prepared from the diluted solutions of surfactin using a negativestaining method, and 1% uranyl acetate solution was used as the staining agent. A drop of the solution was placed on a carbon Formvar-coated copper grid (300 mesh), and the excess liquid was sucked away by filter paper. Then, the sample was negatively stained with 1% uranyl acetate solution. After drying, the samples were imaged under a JEM-200CX electron microscope. Dynamic Light Scattering (DLS). Measurements were carried out at 303.2 ( 0.5 K with an LLS spectrometer (ALV/ SP-125) that employs a multi-τ digital time correlator (ALV5000). A solid-state He-Ne laser (output power of 22 mW at λ ) 632.8 nm) was used as a light source. The samples were introduced into the 7 mL glass bottle through a 0.45 µm filter before measurement. Thereafter, DLS measurements were performed at a scattering angle of 90°. The correlation function of scattering data was analyzed via the CONTIN method to obtain the distribution of diffusion coefficients (D) of the solutes, and then, the apparent equivalent hydrodynamic radius (Rh) was determined using the Stokes-Einstein equation Rh ) kT/6πηD, where k is the Boltzmann constant, T is the absolute temperature, and η is the solvent viscosity. Results and Discussion Micellization of Surfactin. First, we studied the micellization behavior of surfactin in phosphate buffer of pH 7.4 by microcalorimetry. The variation of the observed enthalpy (∆Hobs) with the final surfactin concentration (C) at 303.15 K is presented in Figure 2. ∆Hobs is considerably exothermic at very low surfactin concentration and changes to slightly more exothermic with an increase in surfactin concentration. Beyond 0.038 mM, ∆Hobs begins to change toward less exothermic until reaching an almost horizontal zero value. The critical micelle concentration (CMC) of surfactin can be determined from the intercept of the two lines extrapolated from the initial portion of the curve and the rapidly rising portion of the curve, and the enthalpy of micellization (∆Hmic) can be obtained from the difference between the observed enthalpies of the two linear segments of the plots, as shown in Figure 2. The CMC value of 0.038 mM is relatively low in comparison with that of other ionic surfactants, which shows that surfactin has a much stronger self-assembly ability. Since the positive ∆Hmic observed is not favorable for the micellization process, the entropy should be the main contributor to the surfactin micellization according to enthalpy-entropy compensation theory. Dai and Tam22 concluded that there are two reasons for entropy increase in the surfactant micellization process. On one

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Figure 2. Variation of the observed enthalpy (∆Hobs) with the final surfactin concentration (C) in phosphate buffer of pH 7.4 at 303.15 K.

hand, when hydrophobic segments are dehydrated from an aqueous medium into the surfactant micelle core, the disruption of the solvent cage surrounding the hydrophobic segments in aqueous solutions gives rise to the increase of the entropy. On the other hand, the mobility of the hydrophobic segments increases when the hydrophobes are removed from the aqueous medium to the nonpolar micellar core. Herein, the seven amino acid residues of the surfactin monomer can form strong hydrogen bonds with H2O molecules, and the partial disruption of these hydrogen bonds contributes a great endothermic effect to the surfactin micellization process. That is to say, the favorable entropy increase for surfactin micellization comes not only from the disruption of the solvent cage surrounding the hydrophobic domain of surfactin and the mobility of the hydrophobic parts but also from partial disruption of hydrogen bonds between surfactin molecules and H2O molecules during the micellization process. In order to know the size distribution and morphology of the surfactin aggregates, we performed DLS and TEM measurements. Figure 3 shows the distribution of the hydrodynamic radius for 0.1 and 0.3 mM surfactin solutions expressed by relative intensity and relative number, respectively. In terms of the relative scattered intensity (Figure 3a), the distribution of the hydrodynamic radius is bimodal, with one peak at 4-6 nm and another broad peak centered at ∼85 nm for 0.1 mM surfactin and ∼108 nm for 0.3 mM surfactin. In terms of the relative number (Figure 3b), the distribution of the hydrodynamic radius is only one peak at 4-6 nm for both 0.1 and 0.3 mM surfactin. This demonstrates that the number of small particles at 4-6 nm is actually much more than that of large aggregates. Because the scattering light from the large particles is much stronger than that from the small ones,23 the large particles have a greater effect on the size distribution in terms of relative intensity. The small size of 4-6 nm should be ascribed to the surfactin micelles, whereas the larger aggregates may be the aggregates of small micelles by intermicelle hydrogen bonds. The morphology of surfactin aggregates at 0.3 mM was observed by TEM, as shown in Figure 4, where no vesicles are observed, and proves the coexistence of micelles and large aggregates. Circular dichroism spectra are extremely sensitive to the secondary structure of biomolecules. CD spectra of surfactin have been reported previously.24-27 Ishigami et al.24 found that the surfactin micelles adopted β-sheet conformation in a 0.1 M

Figure 3. Distribution of the hydrodynamic radius for the aggregates of surfactin in pH 7.4 phosphate buffer measured at θ ) 90° and 303.2 ( 0.5 K.

Figure 4. Transmission electron micrograph of 0.3 mM surfactin in pH 7.4 phosphate buffer.

NaHCO3 solution of pH 8.7. Osman et al.25 indicated that both surfactin monomers and surfactin micelles had β-sheet conformation at a neutral pH value, while Maget-Dana et al.26 showed that the surfactin micelles had a turn conformation in pH 8.3 Tris buffer. These results indicate that the secondary structure of surfactin as a peptide is very sensitive to experimental conditions such as electrolytes, pH, and so forth. The β-sheet is the second common form of regular secondary structure in proteins. It consists of β-strands connected laterally by three or more hydrogen bonds. A β-strand is a stretch of typically 5-10 amino acids long, whose peptide backbones are almost fully extended. Such β-strands are arranged adjacent to other strands and form an extensive hydrogen bond network with their neighbors in which the N-H groups in the backbone of one strand establish hydrogen bonds with the CdO groups in the backbone of the adjacent strands, forming a generally twisted, pleated sheet. In general, β-sheet conformation has a minimum at 216-218 nm in the CD spectrum.28 Turn is the shortest and simplest secondary structural element of proteins and plays an important role in the folding of globular proteins, especially for cyclic peptide and oligopeptides.29 Turn conformation falls

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Figure 5. Circular dichroism spectra of surfactin at different concentrations in phosphate buffer of pH 7.4 at 298 ( 1 K.

into β-turn, γ-turn, R-turn, and π-turn according to hydrogen bonding and backbone dihedral angles, in which β-turn is the most common conformation. β-turn in polypeptide chains consists of four amino acids with a hydrogen bond between the CdO group in the first amino acid residue and the N-H group in the fourth amino acid. The typical β-turn CD spectrum has two minima at ∼230 and ∼196 nm plus a maximum at ∼215 nm.30,31 In the present study, far-UV CD spectra of 0.1, 0.3, and 0.5 mM surfactin in 5 mM pH 7.4 phosphate buffer are recorded in Figure 5. The CD spectra exhibit an obvious concentrationdependent manner. The CD spectrum of 0.1 mM surfactin has a negative peak at ∼200 nm and another negative shoulder peak at ∼230 nm. With the surfactin concentration increasing, both minima are blue-shifted, and the corresponding molar ellipticity degree decreases. Until 0.5 mM surfactin, the CD spectrum is characteristic of larger negative peaks at ∼198 and ∼221 nm. Thus, the present results show that the secondary structure of surfactin adopts a β-turn at low micelle concentrations of 0.1 and 0.3 mM and begins to adopt β-sheet conformation at a relatively high micelle concentration of 0.5 mM. By combination of the results from the ITC, DLS, TEM, and CD measurements, we can conclude that surfactin can selfaggregate to micelles at very low concentration and that the micelles tend to aggregate through intermicelle hydrogen bonds. Surfactin can display different secondary structures at different concentrations. It mainly adopts the β-turn conformation at relative low micelle concentration, whereas it adopts the β-sheet conformation at higher micelle concentration. The β-turn may be formed by an intramolecular hydrogen bond, whereas the β-sheet may depend on an intermolecular hydrogen bond. Interaction of Surfactin with Amyloid β(1-40). ITC was performed to monitor the enthalpy change during the interaction of surfactin with Aβ(1-40). The observed enthalpy change of surfactin titrated into 40 µM Aβ(1-40) in pH 7.4 phosphate buffer at 303.15 K is plotted against the final surfactin concentration in Figure 6, together with the curve for the titration of surfactin into the buffer for comparison. The calorimetric curve for the titration of surfactin into Aβ(1-40) shows some difference from that for the titration of surfactin into the buffer. It is obvious that the curve can be divided into three regions. Region I. When the surfactin concentration is lower than 0.038 mM (CMC), the interaction manner of surfactin with Aβ(1-40) starts to change. Since the CMC of surfactin is 0.038 mM, surfactin starts to bind with Aβ(1-40) as micelles beyond 0.038 mM. The micelles continue to bind with Aβ(1-40) through electrostatic attraction between the negative charges of surfactin and the positive charges of Aβ(1-40). The electrostatic binding causes the calorimetric curve of surfactin with Aβ(1-40) to deviate from that of surfactin with buffer, and ∆Hobs becomes more exothermic. At the surfactin concentration of 0.12 mM, the ratio of the positive charges of Aβ(1-40) to the negative charges of surfactin is close to 1, where the positive charges of Aβ(1-40) should have been completely neutralized. Region III. When the surfactin concentration is further increased to above 0.12 mM, the calorimetric curve is almost the same in the presence of Aβ(1-40) as that in the absence of Aβ(1-40). That means that there is no obvious interaction between Aβ(1-40) and surfactin anymore. ∆Hobs of this period only comes from the micelle dilution effect. To understand the interaction process between surfactin and Aβ(1-40), the CD and AFM experiments were conducted to study the structural and conformational variation. The selected surfactin concentrations are 0.01, 0.1, and 0.5 mM, which are located in regions I, II, and III, respectively. The CD spectra and AFM images for Aβ(1-40) in the absence and presence of surfactin in pH 7.4 phosphate buffer are shown in Figures 7 and 8, respectively. As shown by the solid line in Figure 7, Aβ(1-40) alone shows a single minimum at ∼195 nm, which is characteristic of random

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Figure 7. CD spectra of 40 µM Aβ(1-40) itself and in (A) 0.01, (B) 0.1, and (C) 0.5 mM surfactin solution in pH 7.4 phosphate buffer for 2 days. The spectrum of surfactin alone is also illustrated for comparison.

Figure 8. AFM images (2.5 × 2.5 µm2) of 40 µM Aβ(1-40) incubated in (A) 0, (B) 0.01, (C) 0.1, and (D) 0.5 mM surfactin solution in pH 7.4 phosphate buffer for 2 days.

coil conformation.32 From AFM image (Figure 8A), Aβ(1-40) alone displays a larger number of random dispersed globular particles, and the diameter of the particles ranges from 1 to 1.5 nm as determined by the height measurement.

When Aβ(1-40) is incubated in 0.01 mM surfactin solution (region I), the CD spectrum of the mixture exhibits a positive peak at ∼201 nm and a minimum peak at ∼231 nm, which is remarkably different from Aβ(1-40) alone and surfactin alone

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Han et al.

Figure 9. Schematic representation of the possible mechanism for the interactions of Aβ(1-40) with surfactin of different concentration regions.

(Figure 7A). As is well-known, the conversion of soluble Aβ to amyloid fibrils is accompanied by a structural transition from the random coil to β-sheet. The typical β-sheet spectrum has a minimum at around 216 nm and a maximum at about 195 nm in the CD spectra.28 The obtained spectrum in our system is very similar to that of the β-sheet in shape, but it has a red shift of the minimum from ∼216 to ∼231 nm as compared with the normal β-sheet spectrum. This indicates that a large amount of β-sheet conformation does exist, while another conformation also contributes to the whole CD spectrum. As mentioned above, surfactin itself has a special secondary structure in solution, which may affect the CD spectrum of Aβ(1-40). The appearance of the β-sheet signal for region I is due to the presence of a Aβ(1-40) fibrillar structure, which is proved by the AFM image. A very small amount of long fibrils that are 2.5 µm in length and ∼2 nm in diameter is observed in Figure 8B. When Aβ(1-40) is incubated in 0.1 mM surfactin solution (region II), the CD spectrum of the mixture also exhibits a positive peak at ∼201 nm and a minimum peak at ∼231 nm (Figure 7B). However, the amplitude of the ellipticity degree is much larger than that of Aβ(1-40) incubated in 0.01 mM surfactin solution. This shows that Aβ(1-40) has acquired much more β-sheet conformation in 0.1 mM surfactin solution than in 0.01 mM surfactin solution. We indeed observed a lot of fibrils with the diameter of 1.5-4.5 nm and the length ranging from several decades to several hundreds of nanometers from AFM image (Figure 8C), and the amount of the fibrils is significantly much more than that in the presence of 0.01 mM surfactin, which is consistent with the CD results. When Aβ(1-40) is incubated in 0.5 mM surfactin solution (region III), the CD spectrum exhibits a very similar shape to that of 0.5 mM surfactin alone (Figure 7C), except that the amplitude of the band at around ∼200 nm minimum decreases. This shows that the CD signal of the mixture is mainly from the secondary structure of surfactin due to the excessive surfactin as compared with Aβ(1-40), whereas distinct conformational transformation to the β-sheet structure for Aβ(1-40) has not been

observed. The corresponding AFM image exhibits a large amount of globular aggregates with diameters of 2.5-5 nm (Figure 8D), suggesting that Aβ(1-40) is not organized into fibrils. Taking into account the above ITC, AFM, and CD results, the possible mechanism for the interactions of Aβ(1-40) with surfactin of different concentrations is proposed in Figure 9. When the surfactin concentration is below 0.038 mM (region I), surfactin exists as monomers. The surfactin monomers electrostatically bind to positively charge residues of the Aβ(140) peptide chain, and the partial neutralization of positive charges could increase the electrostatic repulsion among the negative charges within the Aβ(1-40) peptide molecule; thus, unordered curled Aβ(1-40) peptide molecules extend. In addition, adjacent Aβ(1-40) molecules already binding with the surfactin monomers may approach each other through hydrophobic interaction between hydrophobic chains of the surfactin monomers, making the conformational transition to the β-sheet structure more easily. When the surfactin concentration falls between 0.038 and 0.12 mM (region II), surfactin starts to form micelles. On one hand, the surfactin micelles bind to the Aβ(140) chain through electrostatic attraction; then, much more net negative charge on the Aβ(1-40) chains leads Aβ(1-40) to unfold to a great extent. On the other hand, the surfactin molecule can form hydrogen bonds with the Aβ(1-40) peptide chain due to the existence of seven amino acid residues in the surfactin molecule. Being driven by the hydrogen bonds, different Aβ(140) peptide chains may further approach each other, thus promoting the conformation transformation of Aβ(1-40) to the β-sheet structure. Additionally, cooperative hydrophobic interaction among the surfactin monomers already bound to Aβ(1-40) may also assist the formation of the β-sheet structure. When the surfactin concentration is beyond 0.12 mM (region III), a large amount of surfactin micelles exists. Since the positive charges of Aβ(1-40) have been saturated by surfactin, the excessive surfactin micelles cannot bind on Aβ(1-40) through electrostatic attraction but separate Aβ(1-40) peptide chains and

Micellization of Surfactin restrict Aβ(1-40) chains from interacting with other Aβ(1-40) peptide chains. Meanwhile, Aβ(1-40) peptide chains are still partially unfolded due to the binding of surfactin micelles, and the complexes of Aβ(1-40) with the surfactin micelles form mixed aggregates through hydrophobic interaction between 29-40 hydrophobic residues of Aβ(1-40) and the surfactin micelles, and also possibly through intermicelle hydrogen bonds of surfactin. Therefore, Aβ(1-40) molecules are inhibited from aggregating into the β-sheet conformation, and fibrils cannot be observed. Conclusions This work investigated the micellization of a biosurfactant surfactin and its effect on the aggregation behavior of Aβ(140). Surfactin can self-aggregate into micelles at very low CMCs, and the micelles tend to aggregate. Surfactin adopts a β-turn conformation at low micelle concentration but the β-sheet at high micelle concentration. The effect of surfactin on the aggregation behavior of Aβ(1-40) shows a strong concentrationdependent fashion. Below CMC, the binding of surfactin monomers on the positive charge residues of Aβ(1-40) increases the electrostatic repulsion among the negative charge residues of Aβ(1-40) and promotes the Aβ(1-40) molecules to unfold. Possibly assisted by the hydrophobic interaction among surfactin monomers on the Aβ(1-40) chain, the conformation of Aβ(140) transfers to the β-sheet structure, which promotes the formation of fibrils. At a low surfactin micelle concentration, the surfactin micelles can interact strongly with Aβ(1-40) to induce a remarkable change of the secondary structure and fibril formation. Besides the electrostatic force and hydrophobic interaction, hydrogen bonds between the surfactin micelles and adjacent Aβ(1-40) chains are the possible driving force to promote the ordered organization of these Aβ(1-40) peptide chains, which leads to the formation of β-sheet and fibrils to a great extent. At relatively high surfactin micelle concentration, the separating of Aβ(1-40) chains by the excessive surfactin micelles and the aggregation of the complexes of Aβ(1-40) with surfactin micelles inhibit the formation of β-sheets and fibrils. Compared with conventional surfactant, surfactin can induce a remarkable secondary structure change and fibril formation of Aβ(1-40) even at a very low concentration (0.01 mM), which shows that surfactin can interact strongly with Aβ(1-40). Due to the existence of seven amino acid residues in the surfactin structure, the hydrogen bond plays an extraordinary role in the interaction of surfactin with Aβ(1-40). This study reveals that the aggregate structure of Aβ(1-40) can be effectively adjusted by a biosurfactant. Acknowledgment. This work is supported by the Chinese Academy of Sciences and the National Natural Science Foundation of China (20633010 and 20573123).

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