Spectrochimica Acta Part A: Molecular and Biomolecular Spectroscopy 124 (2014) 84–90

Contents lists available at ScienceDirect

Spectrochimica Acta Part A: Molecular and Biomolecular Spectroscopy journal homepage: www.elsevier.com/locate/saa

Investigation of interaction of antibacterial drug sulfamethoxazole with human serum albumin by molecular modeling and multi-spectroscopic method Qin Wang ⇑, Sheng-Rui Zhang, Xiaohui Ji School of Chemistry and Environment Science, Shaanxi University of Technology, Hanzhong, Shaanxi 723000, China

h i g h l i g h t s

g r a p h i c a l a b s t r a c t

 The interaction of SMX and HSA was

firstly predicted through molecular modeling.  The binding parameters were performed through the fluorescence quenching spectra.  The thermodynamic parameters were calculated by the Van’t Hoff equations.  The conformational changes of HSA were confirmed by multispectroscopic methods.

a r t i c l e

i n f o

Article history: Received 28 August 2013 Received in revised form 20 December 2013 Accepted 23 December 2013 Available online 7 January 2014 Keywords: Sulfamethoxazole Human serum albumin Molecular interaction Molecular modeling Multi-spectroscopy

a b s t r a c t Interaction of sulfamethoxazole (SMX) with human serum albumin (HSA) was investigated by molecular modeling and multi-spectroscopic methods under physiological conditions. The interaction mechanism was firstly predicted through molecular modeling that confirmed the interaction between SMX and HSA. The binding parameters and the thermodynamic parameters at different temperatures for the reaction had been calculated according to the Stern–Volmer, Hill, Scatchard and the Van’t Hoff equations, respectively. One independent class of binding site existed during the interaction between HSA and SMX. The binding constants decreased with the increasing temperatures, which meant that the quenching mechanism was a static quenching. The thermodynamic parameters of the reaction, namely standard enthalpy DH0 and entropy DS0, had been calculated to be 16.40 kJ mol1 and 32.33 J mol1 K1, respectively, which suggested that the binding process was exothermic, enthalpy driven and spontaneous. SMX bound to HSA was mainly based on electrostatic interaction, but hydrophobic interactions and hydrogen bonds could not be excluded from the binding. The conformational changes of HSA in the presence of SMX were confirmed by the three-dimensional fluorescence spectroscopy, UV–vis absorption spectroscopy and circular dichroism (CD) spectroscopy. CD data suggested that the protein conformation was altered with the reduction of a-helices from 55.37% to 41.97% at molar ratio of SMX/HSA of 4:1. Ó 2014 Elsevier B.V. All rights reserved.

Introduction Sulfonamides that possess a p-aminobenzenesulfonamide framework are a common class of antibiotics and are the first ⇑ Corresponding author. Tel.: +86 916 2642766; fax: +86 916 2641574. E-mail addresses: [email protected] (Q. Wang), [email protected] (S.-R. Zhang). 1386-1425/$ - see front matter Ó 2014 Elsevier B.V. All rights reserved. http://dx.doi.org/10.1016/j.saa.2013.12.100

chemotherapeutic agents effective for the treatment of bacterial and protozoan infections in veterinary and human medicine practices [1–3]. In animal husbandry, sulfonamides are often added to the feed of swine, birds, and cattle for the prevention and treatment of infections or for growth promotion due to the advantages of broad antibacterial spectrum, high efficacy, and low prices [4,5]. Abuse of sulfonamides or insufficient withdrawal time can lead to

Q. Wang et al. / Spectrochimica Acta Part A: Molecular and Biomolecular Spectroscopy 124 (2014) 84–90

accumulation of these drugs in animal tissues, which may cause allergy, carcinogenesis, and formation of resistant bacteria in the human body [6–8]. As a member of the sulfonamide family, sulfamethoxazole (SMX) (structure shown in Fig. 1a) is a now widely used pharmaceutical product in treating urinary tract and lower respiratory tract infections [9–11]. However, SMX could be accumulated in human body and has potential toxicity to human health because of its carcinogenic potency and possible antibiotic resistance [12–14]. This problem has received increasing public attention, and many authorities around the world have proposed maximum residue limits (MRL) of sulfonamides that can be allowed in foods [15,16]. Human serum albumin (HSA) (crystal structure shown in Fig. 1b) is the most abundant protein in plasma and has been the most widely used model protein in evaluating the drug-protein system because it has been extensively characterized [17–19]. HSA contains three homologous domains (I–III): I (residues 1– 195), II (196–383) and III (384–585); each domain can be divided into two subdomains (A and B) [20,21], and there is a large hydrophobic cavity in subdomain IIA where many small molecules can bind [22]. Crystal structure analysis has revealed that amino acid sequence of HSA contains a total of 17 disulfide bridges, one free thiol (Cys-34) and a single tryptophan (Try-214), and according to Peters, the tryptophan residue (Trp214) of HSA is in subdomain IIA [23]. As the major soluble protein constituent of the circulatory system, HSA has many physiological and pharmacological functions which are participating in absorption, distribution, metabolism and excretion of drugs [24–26]. Most drugs travel in plasma and reach the target tissues by binding to HSA, which is an important factor in determining drug pharmacokinetics including distribution and elimination [27–29]. Therefore, studies on the interaction between sulfonamides and HSA are necessary to understand the potential toxicity of sulfonamides to the human bodies. Until now, the mechanisms of the interactions between sulfonamides and HSA have been investigated by several groups. Gao et al. investigated the interaction between HSA and sulfamethazine

85

by capillary electrophoresis, fluorescence spectrometry, and circular dichroism (CD) [30]. Nandibewoor et al. performed the binding of SMX to bovine serum albumin by spectroscopic methods [31]. Zou et al. studied the binding of SMX to HSA by a technique of microdialysis with liquid chromatography [32]. However, the mechanism of the reaction, binding parameters, basic thermodynamic parameters and alteration in the protein secondary structure has not been investigated. In this paper, the mechanism of the reaction was firstly simulated using a SGI FUEL workstation. The binding constants, numbers of binding sites and basic thermodynamic parameters under different temperatures were calculated according to the, Modified Hill equation, Scatchard plots and Van’t Hoff equation. The three dimensional fluorescence spectra were carried out to reveal the changes of HSA’s tryptophan and tyrosine residues and the behavior of HSA’s characteristic polypeptide backbone structure. Meanwhile, the alterations of the protein secondary structures induced by the addition of SMX were further investigated by UV–vis absorption spectroscopy and CD spectroscopy.

Materials and methods Materials SMX and HSA were purchased from Sigma Chemical Company. They were used without further purification. Tris(tris(hydroxymethyl)-amino-methane) was purchased from Tianjin Guangfu Fine Chemical Research Institute (Tianjin, China). All solutions of HSA were prepared in pH 7.40 buffer solution, and stock solutions of HSA (3.0  105 mol L1) were kept in the dark at 4 °C. Tris (0.2 mol L1)-HCl (0.1 mol L1) buffer solution containing NaCl (0.1 mol L1) was used to keep the pH of the solutions at 7.40. NaCl (1.0 mol L1) solution was used to maintain the ionic strength at 0.1. The stock solution (3.0  103 mol L1) of SMX was prepared by dissolving appropriate amounts of sulfonamides in anhydrous methanol and kept at 4 °C. All other reagents were of analytical reagent grade and doubly distilled water was used throughout the experiments. Apparatus All fluorescence spectra were recorded using a F97Pro spectrofluorophotometer (LengGuang Industrial Co., Ltd. of Shanghai, China). Fluorescence emission spectra were recorded from 270 to 450 nm (excitation wavelength 283 nm) using 10 nm/10 nm slit widths. An electronic thermo regulating water-bath (NTT-2100, EYELA, Japan) was used to control the temperature. UV–vis absorption spectra were recorded using a Tu-1901 spectrophotometer (Puxi Analytic Instrument Ltd. of Beijing, China) equipped with a 1.0 cm quartz cell. CD measurements were carried out using an Olis DSM 1000 CD (American) in a cell of path length 1 mm at room temperature. Methods

Fig. 1. (a) The structure of SMX and (b) crystal structure of HSA.

Molecular modeling study The crystal structure of HSA in complex with R-warfarin was taken from the Brookhaven Protein Data Bank (entry codes 1h9z) [33]. The potential of the 3D structure of HSA was assigned based on the Amber 4.0 force field with Kollmanall-atom charges. The initial structure of SMX was generated by molecular modeling software Sybyl 6.9 [34]. The geometry of the molecule was subsequently optimized to minimal energy using the Tripos force field with Gasteiger–Marsili charges, and the FlexX program was

86

Q. Wang et al. / Spectrochimica Acta Part A: Molecular and Biomolecular Spectroscopy 124 (2014) 84–90

used to build the interaction modes between SMX and HSA. All calculations were performed on a SGIFUEL workstation. Spectral measurements Quantitative analysis of the potential interaction between SMX and HSA were carried out by the fluorimetric titration: 3.0 mL solution containing 3.0  106 mol L1 HSA was added to a 1.0 cm quartz cuvette, and then titrated by successive additions of a 3.0  103 mol L1 SMX solution with a trace syringe. These solutions were allowed to stand for 3 min to equilibrate, and the intensities at 332 nm of HSA were recorded at four temperatures (293, 298, 303 and 308 K). The appropriate blanks corresponding to the Tris-HCl buffer solution were subtracted to correct background of fluorescence. The absorption of the molecule at the excitation wavelength or at the emission wavelength will lead to a spurious decrease in the observed fluorescence intensity, and this phenomenon is called the inner-filter effect [35]. During the fluorescence titration experiments, SMX had absorption at the excitation and emission wavelengths of HSA, inducing factitiously reduced emission intensity. Thus, it is very necessary to subtract such effect from the fluorescence quenching data. The extent of inner-filter effect can be eliminated using the following formula [36]:

F c ¼ F o eðAex þAem Þ=2

ð1Þ

where Fc and Fo are the corrected and measured fluorescence, respectively. Aex and Aem are the absorbance of SMX at excitation and emission wavelengths, respectively. The fluorescence intensities used in this paper have been corrected through this formula. Three-dimensional fluorescence spectra were performed under the following conditions: the emission wavelengths were recorded between 220 and 650 nm; the initial excitation wavelengths were set to 220 nm with increment of 5 nm, and other scanning parameters were the same as those of the fluorescence quenching spectra. UV absorbance spectra of the SMX–HSA with concentrations of SMX in the range of 2.0  105–1.0  104 mol L1 were recorded from 190 to 400 nm. CD measurements were performed at room temperature over the wavelengths ranged from 300 to 190 nm. Each spectrum represented the average of five successive scans. Results and discussion Molecular modeling study on the interaction between SMX and HSA The partial binding parameters of the SMX–HSA system were calculated using a SGI FUEL workstation, and the best energy ranked result was shown in Fig. 2. It could be observed that SMX was in the hydrophobic cavity of HSA, and there were two hydrogen bonds between SMX and the amino acid residues: the sulfonyl of SMX donated the O to the H of the arginine residues (Arg222) of HSA. The hydrophobic interaction existed between the leucine residue (Leu238) of HSA and the methyl of SMX. Moreover, the calculated binding Gibbs free energy (DG) was 28.86 kJ mol1, which also indicated the strong interaction between SMX and HSA. These results of molecule modeling provide a theoretical foundation for the following experiments. Fluorescence quenching spectra Fluorescence quenching is the decrease of the quantum yield from a fluorophore induced by a variety of molecular interactions with a quencher molecule, such as excited-state reactions, energy transfer, ground-state complex formation, and collisional quench-

ing [34]. The intrinsic fluorescence of HSA would weaken obviously when the microenvironment of HSA is altered slightly, and the intrinsic fluorescence of HSA is almost contributed by tryptophan residues alone because phenylalanine residues have a very low quantum yield and the fluorescence intensity of tyrosine residues is almost totally quenched when small molecule substances are added [37]. Thus, fluorescence quenching measurements were carried out to investigate the conformational change of HSA, which was shown in Fig. 3. HSA exhibited a strong fluorescence emission with a peak at 332 nm. With gradual increase of SMX concentrations, HSA showed a significant reduction in the fluorescence emission intensity, and a substantial red shift of the maximum emission wavelength from 332 to 338 nm was also observed, which suggested that the polarity of the protein environment was higher compared to that of the pure protein solution. The above phenomenon early indicated that the addition of SMX into HSA changed the microenvironment of tryptophan residues in HSA and the conformational structures of HSA. Binding mechanisms and parameters The quenching mechanisms are usually classified into two different mechanisms: dynamic quenching and static quenching, which can be distinguished by their different dependence on temperature and viscosity or by lifetime measurements [38]. The dynamic quenching results from collision diffusion between the fluorophore and quencher, and higher temperatures will result in larger fluorescence quenching constants [39]. In contrast, the static quenching is caused by the formation of a ground-state complex, and higher temperatures are likely to result in the decreased stability of bound complexes and lower values of the static quenching constants [39]. Dynamic or static quenching can be distinguished by analyzing the fluorescence data with the Stern–Volmer equation

F 0 =F ¼ 1 þ K q s0 ½Q  ¼ 1 þ K SV ½Q 

ð2Þ

where F0 and F are the relative fluorescence intensities of fluorophore in the absence and presence of the quencher, respectively; Kq is the quenching rate constant of the biological macromolecule; s0 is the average lifetime of the molecule without any quencher (for most biomolecules, s0 is about 108 s) [40]; [Q] is the quencher concentration and KSV is the Stern–Volmer quenching constant. The calculated KSV at different temperatures were shown in Table 1. Obviously, as shown in Fig. 4, the Stern–Volmer quenching constant was inversely correlated with temperature. The Kq values decreased from 4.20  1011 to 3.09  1011 L mol1 s1, which was far greater than the maximum scatter collision quenching constant (Kq) of various quenchers with the biopolymer (2  1010 L mol1 s1) [41]. Transparently, the quenching mechanism did not spring from dynamic quenching but proceed from static quenching. For a static quenching interaction, the binding constant (Kb) and the number of binding site (n) can be determined using the Hill equation [42]:

log

F0  F ¼ log K b þ n log½Q  F

ð3Þ

When ligands exist in protein solution and bind partially to sites of protein molecules, free ligand concentration ([Q]free) can be defined as [43]:

½Q free ¼ ½Q   nð1  F=F 0 Þ½P

ð4Þ

where n is the average binding number for one protein molecule, [P] is the total protein concentration, F0, F and [Q] express the same meaning as that in Eq. (2). According to Eq. (4), the Eq. (3) can be described by the following Eq. (5) as follow:

Q. Wang et al. / Spectrochimica Acta Part A: Molecular and Biomolecular Spectroscopy 124 (2014) 84–90

87

Fig. 2. Binding mode between SMX and HSA. The residues of HSA were represented using lines, the SMX structure was represented using ball and stick model and hydrogen bonds between SMX and HSA were represented using dashed lines.

Fig. 3. The fluorescence spectra of SMX–HSA system. (a) HSA; (b–f) SMX–HSA; [HSA] = 3.0  106 mol L1, [SMX] = 2.0  105–10.0  105 mol L1, (g) 5.0  105 mol L1 SMX; Tris buffer (pH 7.40), T = 293 K.

Table 1 The Stern–Volmer and modified Hill equation quenching parameters for SMX–HSA system at different temperatures. Temperature (K)

293 298 303 308

log

The binding constants and the numbers of binding sites were also calculated using the Scatchard equation to confirm how many molecules of the SMX bound to HSA.

r=Df ¼ nK  rK

Stern–Volmer

Modified Hill equation

KSV (103 L mol1)

Kb (103 L mol1)

n

4.20 3.64 3.29 3.09

4.28 3.55 3.35 3.17

1.00 1.00 1.00 1.00

F0  F ¼ log K b þ n log f½Q   ð1  F=F 0 Þ½Pg F

Fig. 4. Stern–Volmer plots of fluorescence quenching of the SMX–HSA system at four different temperatures (j 293 K,  298 K, N 303 K, . 308 K). [HSA] = 3.0  106 mol L1, [SMX] = 0.0  105–10.0  105 mol L1, Tris buffer (pH 7.40); kex = 283 nm, kem = 332 nm.

ð5Þ

The binding constant (Kb) and number of equivalent binding sites (n) could be determined, which were shown in Fig. 5. According to a plot of log [(F0F)/F] versus log [Q], the binding constants and the numbers of binding sites were summarized in Table 1. The values of n for HSA were approximately equal to 1, indicating that there was one binding site in HSA for SMX during their interaction.

ð6Þ

where r is the number of moles of ligand bound per mole of protein; Df is the molar concentration of free ligand; n and K are the number of binding site and binding constant, respectively. The binding constants and numbers of binding sites were the slopes and the slopes divided by intercepts, respectively, which were presented in Fig. 6 and Table 2. There was linear dependence between r/Df and r, and the slopes decreased with the increase of temperatures, which indicated that the quenching was a static quenching process. Meanwhile, the linearity of Scatchard plots indicated that the SMX bound to one class of site on HSA, which was consistent with the conclusion obtained from Hill equation (n = 1). Binding modes In general, the interaction forces between small molecular substrates and biological macromolecules were four representative

88

Q. Wang et al. / Spectrochimica Acta Part A: Molecular and Biomolecular Spectroscopy 124 (2014) 84–90

ln K ¼ DH=RT þ DS=R

ð7Þ

K is the binding constant at the corresponding temperature; R and T are the gas constant and absolute temperature, respectively; DH and DS are enthalpy change and entropy change, respectively. The free energy change DG is estimated from the equation:

DG ¼ DH  T DS

Fig. 5. Modified Hill equation plots of fluorescence quenching of the SMX–HSA system at four different temperatures (j 293 K,  298 K, N 303 K, . 308 K). [HSA] = 3.0  106 mol L1, [SMX] = 2.0  105–10.0  105 mol L1, Tris buffer (pH 7.40); kex = 283 nm, kem = 332 nm.

ð8Þ

According to the binding constants of SMX to HSA, DH was calculated from the slope of the Van’t Hoff relationship; DS was calculated from the intercept (shown in Fig. 7), and all the results were presented in Table 2. DH for the binding reactions between SMX and HSA was 16.40 kJ mol1, and DS for the binding reactions between SMX and HSA was 32.33 J mol1 K1. The negative sign for DH indicated that the formation of the SMX–HSA complex was exothermic, and the positive sign for DS characterized that the binding process was entropy-driven. The negative sign for DG manifested that the binding interaction was spontaneous. From the point of view of water structure, negative enthalpy change is generally considered as a typical evidence for electrostatic interaction, and positive entropy change arises from hydrophobic and electrostatic interaction. In this paper, negative DH and positive DS values indicated that the electrostatic interactions played major roles in the SMX–HSA binding reaction, and because of the positive sign for DS, the hydrophobic interaction was also the possible force of interaction between SMX and HSA. Effect of SMX on the HSA conformations

Fig. 6. Scatchard equation plots of fluorescence quenching of the SMX–HSA system at four different temperatures (j 293 K,  298 K, N 303 K, . 308 K). [HSA] = 3.0  106 mol L1, [SMX] = 2.0  105–10.0  105 mol L1, Tris buffer (pH 7.40); kex = 283 nm, kem = 332 nm.

Table 2 The Scatchard equation quenching parameters and thermodynamic parameters for SMX–HSA system at different temperatures. Temperature (K)

Scatchard equation DG (kJ mol1) K n 3 1 (10 L mol )

293 298 303 308

4.11 3.72 3.23 2.99

1.02 0.99 1.01 1.02

25.87 26.03 26.20 26.36

DS (J mol1 K1)

DH (kJ mol1)

32.33

16.40

types, including hydrophobic force, hydrogen bond, van der Waals force, and electrostatic interaction. As demonstrated by Ross and Subramanian, the thermodynamic parameters of the reactions comprise the major evidences that confirm the binding modes, and the signs and magnitudes of the enthalpy (DH) and entropy (DS) can account for the main forces contributing to protein stability [44]. In this paper, the binding constants obtained from Scatchard equations were applied to the discussion of binding modes and the main forces. The values of the enthalpy (DH) and entropy (DS) dependent on temperatures can be calculated by Van’t Hoff equation as follows:

Recently, three-dimensional fluorescence spectroscopy as a newly developed fluorescence analytical technique is used to investigate the conformation change of HSA more carefully. As shown in Fig. 8, the three-dimensional fluorescence spectra of HSA in absence and presence of SMX were investigated, and the related characteristic parameters were listed in Table 3. The peak 1 at the left was the first-ordered Rayleigh scattering peak (kex = kem), whereas the peak 3 at the right was the second-ordered Rayleigh scattering peak (kem = 2kex) [33]. The intensity of peak 1 and peak 3 both decreased with the addition of SMX. The reason of the phenomenon may be attributed to the formation of the SMX–HSA complex that leaded to the increase of the macromolecule diameter. In addition to peak 1 and peak 3, peak 2 as one strong fluorescence peak revealed the spectral characteristics of tryptophan and tyrosine residues and the fluorescence spectral behavior of polypeptide backbone structures whose intensity was correlated with the secondary structure of protein [45]. As shown in Fig. 8 and Table 3, the addition of SMX to HSA leaded to a significant

Fig. 7. Van’t Hoff plots of the SMX–HSA system.

Q. Wang et al. / Spectrochimica Acta Part A: Molecular and Biomolecular Spectroscopy 124 (2014) 84–90

89

Fig. 8. Three-dimensional fluorescence spectra of the free HSA (a) and SMX–HSA system (b). The concentrations of HSA were both 3.0  106 mol L1; [SMX]: (a) 0.0 mol L1; (b) 6.0  105 mol L1; T = 293 K.

Table 3 Three-dimensional fluorescence spectral characteristics of HSA and SMX–HSA systems. System

Peak 2 (kex/kem)

Dk (nm)

Intensity

HSA

283/332 283/338

49 55

513.682 381.573

20:1

reduction in the fluorescence intensities with a red shift of maximum emission wavelength shift from 332 to 338. It was concluded that the interaction of SMX with HSA induced the slight conformational change of the protein. The UV–vis absorption spectra of HSA and SMX–HSA system were carried out to confirm the quenching mechanism and conformational changes of HSA caused by the addition of SMX [46]. As the spectra were shown in Fig. 9, the absorbance of HSA at 213 nm decreased with the addition of SMX, and red shift of the spectra in SMX–HSA system from 213 to 219 nm could be observed. It is well accepted that the dynamic quenching is largely caused by the collision and the energy transfer from HSA to drugs in this way, which would not cause the UV spectrum change of HSA. Otherwise, a HSA–drug complex formed in the static quenching would induce the changes of the UV spectrum of HSA. Therefore, the fluorescence quenching of HSA in our case was primarily caused by complex formation between SMX and HSA, which was accordance with the fluorescence quenching measurement. Furthermore, the absorption of HSA (about 210 nm) is the characteristic of a-helical structure of HSA, which clearly indicated that the interaction between SMX and HSA induced the change in a-helix structure of the protein. Fig. 10 shows the CD spectra of HSA with various concentrations of the SMX, which exhibited characteristic features of the typical

Fig. 9. UV–vis absorption spectra of the SMX–HSA system. (a) HSA; (b–f) SMX– HSA; [HSA] = 3.0  106 mol L1, [SMX] = 2.0  105–10.0  105 mol L1; Tris buffer (pH 7.40), T = 293 K.

(a + b) helix structure of the free HSA and its SMX complex with negative bands at 208 and 218 nm [47]. The binding of SMX to HSA caused a significant intensity decrease for both negative bands, indicating a decrease in the a-helical content of the protein. From the above results, it could be seen that the addition of SMX to HSA caused a conformational change in the protein, and the a-helical contents of HSA in the absence and presence of SMX were calculated by quantitative analysis of the CD spectra data. The induced ellipticities were obtained by subtracting the ellipticities of SMX from the ellipticities of the SMX–HSA mixtures at the same wavelength and were expressed in millidegree. Results were expressed as mean residue ellipticity (MRE) in deg cm2 dmol1, which was defined by the following equation:

90

Q. Wang et al. / Spectrochimica Acta Part A: Molecular and Biomolecular Spectroscopy 124 (2014) 84–90

Fig. 10. CD spectra of the SMX–HSA system. The concentration of HSA was 3.0  106 mol L1 while the SMX concentration was 12.0  106 mol L1; T = 293 K.

MRE ¼ hobs ðm degÞ=10nlC p

ð9Þ

hobs represents the CD in millidegree, n is the number of amino acid residues, l is the path length of the cell and Cp is the mole fraction. The a-helical contents of HSA were calculated from the molar ellipticities ([h]) at 208 nm using the following equation:





a-helixð%Þ ¼ ð½h208  4000Þ=ð33; 000  4000Þ  100

ð10Þ

From the above equation, the calculating results exhibited that

a-helix structures of HSA decreased from 55.37% to 41.97% at molar ratio of SMX/HSA of 4:1, which was indicative of the loss of a-helical content. Conclusions The above research work provided an approach for studying interaction of sulfonamides with HSA by molecular modeling and multi-spectroscopic techniques. The results of molecule modeling performed that the strong interaction existed between SMX and HSA, which provided a theoretical foundation for the following experiments. The experimental results from the fluorescence quenching indicated the presence of static quenching mechanism in the binding of SMX to HSA. The binding of SMX to HSA was found to be spontaneous, while the hydrophobic interactions and electrostatic forces played a major role in the interaction. The three dimensional fluorescence spectroscopy, UV–vis absorption spectroscopy and CD spectroscopy data revealed the conformational changes of HSA upon its interaction with SMX. The work presented here can be helpful for evaluation of side effects of sulfonamides like SMX and provide a theoretical basis for further researches on other drugs. Acknowledgment The authors acknowledge the financial support from the startup project of Shaanxi University of Technology (SLGQD133). References [1] Y.C. Guo, B. Ngom, T. Le, X. Jin, L.P. Wang, D.S. Shi, X.L. Wang, D. Bi, Anal. Chem. 82 (2010) 7550–7555.

[2] B.B. Zhang, Z.G. Chen, Y.Y. Yu, J.P. Yang, J.B. Pan, Chromatographia 76 (2013) 821–829. [3] M.J. García-Galán, M.S. Díaz-Cruz, D. Barceló, Talanta 81 (2010) 355–366. [4] L.L. Wang, J. Wu, Q. Wang, C.H. He, L. Zhou, J. Wang, Q.S. Pu, J. Agric. Food Chem. 60 (2012) 1613–1618. [5] S.L. Borràs, R. Companyó, J. Guiteras, J. Agric. Food Chem. 59 (2011) 5240– 5247. [6] L.G. Chen, X.P. Zhang, L. Sun, Y. Xu, Q.L. Zeng, H. Wang, H.Y. Xu, A.M. Yu, H.Q. Zhang, L. Ding, J. Agric. Food Chem. 57 (2009) 10073–10080. [7] B.Y. Liu, X.P. Nie, W.Q. Liu, P. Snoeijs, C. Guan, M.T. Tsui, Ecotoxicol. Environ. Saf. 74 (2011) 1027–1035. [8] J. Bernal, M.J. Nozal, J.J. Jiménez, M.T. Martín, E. Sanz, J. Chromatogr. A 1216 (2009) 7275–7280. [9] X.P. Nie, B.Y. Liu, H.J. Yu, W.Q. Liu, Y.F. Yang, Environ. Pollut. 172 (2013) 23–32. [10] A. Dirany, S.E. Aaron, N. Oturan, I. Sirés, M. Oturan, J. Aaron, Anal. Bioanal. Chem. 400 (2011) 353–360. [11] L.L. Ji, Y.Q. Wan, S.R. Zheng, D.Q. Zhu, Environ. Sci. Technol. 45 (2011) 5580– 5586. [12] I.S. Silva, D.T.R. Vidal, C.L. do Lago, L. Angnes, J. Sep. Sci. 36 (2013) 1405–1409. [13] L.S. Andrade, R.C. Rocha-Filho, Q.B. Cass, O. Fatibello-Filho, Anal. Methods 2 (2010) 402–407. [14] N. Sun, S.L. Wu, H.X. Chen, D.J. Zheng, J.W. Xu, Y. Ye, Microchim. Acta 179 (2012) 33–40. [15] S. Lamba, S.K. Sanghi, A. Asthana, M. Shelke, Anal. Chim. Acta 552 (2005) 110– 115. [16] C. Hartig, T. Storm, M. Jekel, J. Chromatogr. A 854 (1999) 163–173. [17] W.H. Gao, N.N. Li, G.P. Chen, Y.P. Xu, Y.W. Chen, S.L. Hu, Z.D. Hu, J. Lumin. 131 (2011) 2063–2071. [18] S. Tabassum, W.M. Al-Asbahy, M. Afzal, F. Arjmand, J. Photochem. Photobiol. B 114 (2012) 132–139. [19] Y.J. Hu, Y.O. Yang, C.M. Dai, Y. Liu, X.H. Xiao, Biomacromolecules 11 (2009) 106–112. [20] U. Kragh-Hansen, Pharmacol. Rev. 33 (1981) 17–53. [21] I. Sjöholm, B. Ekman, A. Kober, I. Ljungstedt-Påhlman, B. Seiving, T. Sjödin, Mol. Pharmacol. 16 (1979) 767–777. [22] X.M. He, D.C. Carter, Nature 358 (1992) 209–215. [23] T. Peters, Biochemistry, Genetics and Medical Applications, Academic Press, San Diego, CA, 1996. [24] Y.J. Hu, Y. Liu, Z.B. Pi, S.S. Qu, Bioorg. Med. Chem. 13 (2005) 6609–6614. [25] X.L. Han, F.F. Tian, Y.S. Ge, F.L. Jiang, L. Lai, D.W. Li, Q.L. Yu, J. Wang, C. Lin, Y. Liu, J. Photochem. Photobiol. B 109 (2012) 1–11. [26] R. Punith, A.H. Hegde, S. Jaldappagari, J. Fluoresc. 21 (2011) 487–495. [27] M.R. Housaindokht, Z. Rouhbakhsh Zaeri, M. Bahrololoom, J. Chamani, M.R. Bozorgmehr, Spectrochim. Acta A 85 (2012) 79–84. [28] K. Joseph, D.S. Hage, J. Chromatogr. B 878 (2010) 1590–1598. [29] C. Brauckmann, C. Wehe, M. Kieshauer, C. Lanvers-Kaminsky, M. Sperling, U. Karst, Anal. Bioanal. Chem. 405 (2013) 1855–1864. [30] J.B. Chen, X.F. Zhou, Y.L. Zhang, H.P. Gao, Sci. Total Environ. 432 (2012) 269– 274. [31] P. Naik, S. Chimatadar, S. Nandibewoor, Spectrochim. Acta A 73 (2009) 841– 845. [32] H.L. Wang, H.F. Zou, A.S. Feng, Y.K. Zhang, Anal. Chim. Acta 342 (1997) 159– 165. [33] X.M. Zhou, Q. Yang, X.Y. Xie, Q. Hu, F.M. Qi, Z.U. Rahman, X.G. Chen, Dyes Pigments 92 (2012) 1100–1107. [34] X.R. Wang, X.Y. Xie, C.L. Ren, Y. Yang, X.M. Xu, X.G. Chen, Food Chem. 127 (2011) 705–710. [35] X.H. Wu, J.J. Liu, H.M. Huang, W.W. Xue, X.J. Yao, J. Jin, Int. J. Biol. Macromol. 49 (2011) 343–350. [36] G.W. Zhang, Y.D. Ma, Food Chem. 136 (2012) 442–449. [37] J. Ma, Y. Liu, L. Chen, Y. Xie, L.Y. Wang, M.X. Xie, Food Chem. 132 (2012) 663– 670. [38] J.R. Lakowicz, Principles of Fluorescence Spectroscopy, third ed., Springer, New York, 2006. [39] T.C. Xu, X.J. Guo, L. Zhang, F. Pan, J. Lv, Y.Y. Zhang, H.J. Jin, Food Chem. Toxicol. 50 (2012) 2540–2546. [40] A. Papadopoulou, R.J. Green, R.A. Frazier, J. Agric. Food Chem. 53 (2005) 158– 163. [41] C. Qin, M.X. Xie, Y. Liu, Biomacromolecules 8 (2007) 2182–2189. [42] J. Min, X.M. Xia, Z. Dong, L. Yuan, L.X. Yu, C. Xing, J. Mol. Struct. 692 (2004) 71– 80. [43] D.W. Lu, X.C. Zhao, Y.C. Zhao, B.C. Zhang, B. Zhang, M.Y. Geng, R.T. Liu, Food Chem. Toxicol. 49 (2011) 3158–3164. [44] P.D. Ross, S. Subramanian, Biochemistry 20 (1981) 3096–3102. [45] Y.Z. Zhang, B. Zhou, X.P. Zhang, P. Huang, C.H. Li, Y. Liu, J. Hazard. Mater. 163 (2009) 1345–1352. [46] X.R. Pan, P.F. Qin, R.T. Liu, J. Wang, J. Agric. Food Chem. 59 (2011) 6650–6656. [47] L.B. Chen, M.H. Wu, X.C. Lin, Z.H. Xie, Luminescence 26 (2011) 172–177.

Investigation of interaction of antibacterial drug sulfamethoxazole with human serum albumin by molecular modeling and multi-spectroscopic method.

Interaction of sulfamethoxazole (SMX) with human serum albumin (HSA) was investigated by molecular modeling and multi-spectroscopic methods under phys...
2MB Sizes 0 Downloads 0 Views