Investigation of the binding of Salvianolic acid B to human serum albumin and the effect of metal ions on the binding

The studies on the interaction between HSA and drugs have been an interesting research field in life science, chemistry and clinical medicine. There are also many metal ions present in blood plasma, thus the research about the effect of metal ions on the interaction between drugs and plasma proteins is crucial. In this study, the interaction of Salvianolic acid B (Sal B) with human serum albumin (HSA) was investigated by the steady-state, synchronous fluorescence and circular dichroism (CD) spectroscopies. The results showed that Sal B had a strong ability to quench the intrinsic fluorescence of HSA through a static quenching mechanism. Binding parameters calculated showed that Sal B was bound to HSA with the binding affinities of 10(5) L mol(-1). The thermodynamic parameters studies revealed that the binding was characterized by positive enthalpy and positive entropy changes, and hydrophobic interactions were the predominant intermolecular forces to stabilize the complex. The specific binding distance r (2.93 nm) between donor (HSA) and acceptor (Sal B) was obtained according to F?rster non-radiative resonance energy transfer theory. The synchronous fluorescence experiment revealed that Sal B cannot lead to the microenvironmental changes around the Tyr and Trp residues of HSA, and the binding site of Sal B on HSA is located in hydrophobic cavity of subdomain IIA. The CD spectroscopy indicated the secondary structure of HSA is not changed in the presence of Sal B. Furthermore, The effect of metal ions (e.g. Zn(2+), Cu(2+), Co(2+), Ni(2+), Fe(3+)) on the binding constant of Sal B-HSA complex was also discussed.     

Studies on the interaction between docetaxel and human hemoglobin by spectroscopic analysis and molecular docking

The binding reaction between docetaxel (DTX) and human hemoglobin (HHb) was investigated systematically with various spectroscopic methods including fluorescence quenching technique, ultraviolet (UV)-vis absorption, synchronous fluorescence, circular dichroism (CD) and Fourier transform infrared (FT-IR) spectroscopy. Analysis of fluorescence data showed that the quenching mechanism was the dynamic quenching and each protein had only one binding site for the drug. Two thermodynamic parameters, the enthalpy change and the entropy change were calculated to be 9.18 kJ mol(-1) and 116J mol(-1) K(-1), respectively, which suggested that hydrophobic interaction played a major role in the binding reaction. The results from different spectroscopic methods also showed that DTX could induce conformational changes of HHb. The molecular docking simulation demonstrated that DTX was located in the central cavity of HHb.     

Investigation of the interaction between human hemoglobin and five antioxidants by fluorescence spectroscopy and molecular modeling

L-ascorbic acid, α-tocopherol, procyanidin B3, β-carotene, and astaxanthin are five classic dietary antioxidants. In this study, the interaction between the five antioxidants and human hemoglobin (HHb) was investigated by fluorescence spectroscopy and molecular modeling. The quenching mechanisms of HHb by the five antioxidants are all static quenching. The downward curvature of the Stern–Volmer plots for HHb–procyanidin B3 system at higher concentrations of procyanidin B3 come from the reason for the variation in the number of accessible tryptophan (Trp) residues toward HHb. The upward curvature of the Stern–Volmer plots for HHb–β-carotene system at higher concentrations of β-carotene predominantly by the “sphere of action” quenching mechanism. The binding constants of HHb with the five antioxidants are in the following order as: astaxanthin > L-ascorbic acid > β-carotene > α-tocopherol > procyanidin B3 at 298 K. The binding processes of the five antioxidants to HHb are all entropy process. Thermodynamic analysis and molecular modeling suggest that the hydrophobic forces are the main interaction force in the binding of the five antioxidants to HHb and hydrogen bond interactions between HHb and L-ascorbic acid/α-tocopherol/procyanidin B3/astaxanthin should be also considered. The fluorescence experimental results are in agreement with the results obtained by molecular modeling study.     

Multi-spectroscopic Investigations of Aspirin and Colchicine Interactions with Human Hemoglobin: Binary and Ternary Systems

The interactions of colchicine (COL) and aspirin (ASA) with human hemoglobin (HB) was studied by fluorescence, UV/vis absorption, resonance light scattering, synchronous fluorescence and circular dichroism (CD) spectroscopic techniques under physiological conditions. The inherent binding information, including the quenching mechanism, binding constants, number of binding sites, effective quenching constant, fraction of the initial fluorescence and thermodynamic parameters were determined by the fluorescence quenching technique at different temperatures. The results proved that the mechanism of fluorescence quenching of HB by COL and ASA is due to formation of HB–drug complexes in the binary and ternary systems. The distance between the acceptor drugs and HB was estimated by Förster’s equation on the basis of fluorescence energy transfer. In addition, according to the synchronous fluorescence spectra of HB, the results showed that the fluorescence quenching of HB originated solely from the tryptophan residues and indicated a conformational change for HB caused by addition of the drugs. Far-UV CD spectra of HB were recorded before and after the addition of ASA and COL both as binary and ternary systems. An increase in intensity of the positive CD peak of HB was observed in the presence of these drugs. The results were interpreted as excited state interactions between the aromatic residues of the HB binding sites and the drugs bound to them.     

Interaction Between Fluorinated Amphiphilic Copolymer P(HFMA)-g-P(SPEG) and BSA

A novel fluorinated amphiphilic copolymer P(HFMA)-g-P(SPEG) was synthesized. The interactions between P(HFMA)-g-P(SPEG) and bovine serum albumin (BSA) were studied by synchronous fluorescence and intrinsic fluorescence spectroscopy. It was concluded through synchronous fluorescence that P(HFMA)-g-P(SPEG) mainly bound to tryptophan residues of BSA. Intrinsic fluorescence results revealed that BSA and P(HFMA)-g-P(SPEG) had strong interactions. The mechanism of quenching belonged to dynamic quenching and the main sort of binding force was hydrophobic force. The hydrophobic interaction between P(HFMA)-g-P(SPEG) and BSA was conformed by micropolarity and TEM photographs.     

Effect of pH on the interaction of vitamin B12 with bovine serum albumin by spectroscopic approaches

The interaction mechanism between vitamin B12 (B12, cyanocobalamin) and bovine serum albumin (BSA) has been investigated by fluorescence, synchronous fluorescence, ultraviolet-vis (UV) absorbance, and three-dimensional fluorescence. The intrinsic fluorescence of BSA was strongly quenched by the addition of B12 in different pH buffer solutions (pH 2.5, 3.5, 5.0, 7.4, and 9.0) and spectroscopic observations are mainly rationalized in terms of a static quenching process at lower concentration of B12 (C(B12)/C(BSA)<5) and a combined quenching process at higher concentration of B12 (C(B12)/C(BSA)>5). The structural characteristics of B12 and BSA were probed, and their binding affinities were determined under different pH conditions. The results indicated that the binding abilities of B12 to BSA in the acidic and basic pH regions (pH 2.5, 3.5, 5.0, and 9.0) were lower than that at simulating physiological condition (pH 7.4). In addition, the efficiency of energy transfer from tryptophan fluorescence to B12 was found to depend on the binding distance r between the donor and acceptor calculated using F?rster's theory. The effect of B12 on the conformation of BSA was analyzed using UV, synchronous fluorescence and three-dimensional fluorescence under different pH conditions. These results showed that the binding of B12 to BSA causes apparent change in the secondary and tertiary structures of BSA.     

Influence of the methyl position on the binding of 5-epi-taiwaniaquinone G to HHb investigated by spectrofluorimetry and molecular modelling

The interactions of human haemoglobin (HHb) with 5-epi-taiwaniaquinone G (G1) and one structural analogue of 5-epi-taiwaniaquinone G (G2) systematically by UV–vis absorption spectroscopy and fluorescence spectroscopy in combination with molecular modelling are studied in this paper. They caused the fluorescence quenching of HHb by the formation of complex. The binding constants and thermodynamic parameters were obtained. The hydrophobic and electrostatic interactions were the predominant intermolecular forces to stabilise these complexes. Results of thermodynamic analysis and molecular modelling showed that G2 was the stronger quencher and bound to HHb with higher affinity. The result of molecular modelling is close to that obtained by experimental methods.     

光谱法研究Cu2+与肌红蛋白的相互作用

用紫外吸收光谱、荧光光谱、同步荧光光谱及圆二色(CD)谱研究了Cu2+与肌红蛋白(Mb)的相互作用. 结果发现, Cu2+使Mb的紫外吸收增强, 峰位蓝移, 说明Cu2+与Mb发生了较强的相互作用; Mb的特征荧光峰猝灭, 且随着温度升高猝灭常数Ksv降低, 表明Cu2+对Mb的荧光猝灭机制属于静态猝灭; 计算了不同温度下的结合常数和结合位点数; 由van′t Hoff方程计算出ΔH和ΔS分别为-11.60 kJ/mol和33.77 J·(mol·K)-1, 得出二者之间的作用力主要为静电力; 并依据Förster非辐射能量转移理论确定了给体-受体间的结合距离r=2.56 nm. 同步荧光光谱表明, Cu2+对Mb的构象产生影响, 使色氨酸残基的疏水性下降. CD光谱测得加入Cu2+后, 二级结构发生改变, 使α-螺旋含量降低.     

铱(IV)离子与人血丙种球蛋白的作用研究

在0.1 mol•L^－1醋酸-醋酸钠(pH 5.0)体系中, 采用紫外吸收光谱、荧光光谱及同步荧光光谱法研究了人血丙种球蛋白(gamma seroglobulinum humanum, 简称GSH)与铱(IV)离子的相互作用. 结果表明, Ir(IV)离子使人血丙种球蛋白的构象发生了改变, α-螺旋含量减少, 并且用同步荧光光谱发现Ir(IV)离子与人血丙种球蛋白的作用位点更接近于色氨酸, 从而使色氨酸残基的疏水性略有减小. 荧光光谱结果表明Ir(IV)对人血丙种球蛋白内源荧光(342 nm)产生了较强的荧光猝灭作用, 根据不同温度下Ir(IV)对人血丙种球蛋白的荧光猝灭作用, 证明了这种荧光猝灭为静态猝灭机制, 计算了其结合常数和结合位点数, 从而得出了静电作用力为其主要的作用力.     

铜(Ⅱ)离子与神经红蛋白的相互作用

利用紫外可见吸收光谱、荧光光谱、同步荧光光谱及圆二色(CD)光谱研究了铜髤离子与神经红蛋白(NGB)的相互作用。结果表明,Cu2+离子使NGB在280nm处的紫外吸收增强,说明Cu2+与NGB发生了相互作用;Cu2+使NGB内源性荧光发生猝灭,其猝灭机制为静态猝灭;同步荧光光谱表明,Cu2+使色氨酸微环境的疏水性有所降低,Cu2+对NGB的作用位点更接近于色氨酸;CD光谱显示Cu2+没有引起NGB二级结构明显的变化。     

Interactions of lysozyme with 6-amino-4-aryl-3-methyl-1-phenyl-1H-pyrazolo[3,4-b]pyridine-5-carbonitriles: a fluorescence quenching study.

The interactions between 6-amino-4-aryl-3-methyl-1-phenyl-1H-pyrazolo[3,4-b]pyridine-5-carbonitrile and lysozyme (LYSO) were investigated by using tryptophane fluorescence quenching and 6-amino-4-(2-hydroxyphenyl)-3-methyl-1-phenyl-1H-pyrazolo[3,4-b]pyridine-5-carbonitrile (1) was studied in detail because of its high water solubility. At different temperatures, the quenching constants K(SV), the binding constants K and the binding sites n of LYSO with 1 were determined and the thermodynamic parameters were calculated. The distance r between tryptophane residues and 1 was obtained according to the Forster mechanism of non-radiation energy transfer. Furthermore, synchronous fluorescence spectroscopy data indicated that the association between 1 and LYSO changed LYSO's conformation and that the hydrophobic interaction played a major role in 1-LYSO association. It was proved that the fluorescence quenching of LYSO by 1 was related to the formation of a 1-LYSO complex and to a non-radiation energy transfer.     

Spectroscopic studies on the interaction between human hemoglobin and CdS quantum dots

The interaction between human adult hemoglobin (Hb) and bare CdS quantum dots (QDs) was investigated by fluorescence, synchronous fluorescence, circular dichroism (CD), and Raman spectroscopic techniques under physiological pH 7.43. The intrinsic fluorescence of Hb is statically quenched by CdS QDs. The quenching obeys the Stern-Volmer equation, with an order of magnitude of binding constant (K) of 10(7). The electrostatic adsorption of Hb on the cationic CdS QDs surface is energetically favorable (DeltaS(0)=70.22 Jmol(-1)K(-1), DeltaH(0)=-23.11 kJmol(-1)). The red shift of synchronous fluorescence spectra revealed that the microenvironments of tryptophan and tyrosine residues at the alpha(1)beta(2) interface of Hb are disturbed by CdS QDs, which are induced from hydrophobic cavities to a more exposed or hydrophilic surrounding. The secondary structure of the adsorbed Hb has a loose or extended conformation for which the content of alpha-helix has decreased from 72.5 to 60.8%. Moreover, Raman spectra results indicated that the sulfur atoms of the cysteine residues form direct chemical bonds on the surface of the CdS QDs. The binding does not significantly affect the spin state of the heme iron, and deoxidation is not expected to take place on the coated oxyhemoglobin. The change of orientation of heme vinyl groups was also detected.     

Interaction of tetrandrine with human serum albumin: a fluorescence quenching study.

The interaction of tetrandrine with human serum albumin (HSA) was studied by measuring fluorescence quenching spectra, synchronous fluorescence spectra and ultra-violet spectra. The fluorescence quenching spectra of HSA in the presence of tetrandrine showed that tetrandrine quenched the fluorescence of HSA. The quenching constants of tetrandrine on HSA were determined using the Stern-Volmer equation. Static quenching and non-radiation energy transfer were the two main reasons leading to the fluorescence quenching of HSA by tetrandrine. According to the F?rster theory of non-radiation energy transfer, the binding distances (r) and the binding constants (K(A)) were obtained. The thermodynamic parameters obtained in this study revealed that the interaction between tetrandrine and HSA was mainly driven by a hydrophobic force. The conformational changes of HSA were investigated by synchronous spectrum studies.     

Interaction of Clofazimine with Divalent Metal Ions: A Fluorescence Quenching Study

The interaction of clofazimine (CFZ) with divalent metal cations was studied by measuring fluorescence quenching spectra, synchronous fluorescence spectra and ultraviolet spectra. The fluorescence quenching spectra of CFZ in the presence of metal ions showed that metal ions quenched the fluorescence of CFZ. The quenching constants were determined using the Stern–Volmer equation. The binding constants (log K) and binding site were obtained. The thermodynamic parameters obtained in this study revealed that the interaction between CFZ and metal ions was mainly driven by a hydrophobic force. The conformational changes of CFZ were investigated by synchronous spectrum studies.     

分子内电荷转移荧光探针1-酮-2-（对二甲氨基苯亚甲基）四氢萘与人血清白蛋白作用

采用荧光及紫外光谱研究了1-酮-2-（对二甲氨基苯亚甲基）-四氢萘（KDTN）与人血清白蛋白（HSA）相互作用的光谱特性。 结果表明,静态猝灭和非辐射能量转移是导致KDTN对HSA荧光猝灭的主要原因。 测得17、27和37 ℃ 3个温度下的结合常数K_A分别为1.633×10⁸、0.7998×10⁸和0.347×10⁸ L/mol,结合位点数n分别为1.7、1.6和1.7;据Forster偶极 偶极非辐射能量转移理论,计算得到KDTN与HSA在3个温度下的作用距离r分别为2.64、2.59和2.64 nm;能量转移效率E分别为0.5100、0.4797和0.4210。 热力学参数表明,二者主要以范德华力或氢键结合;用同步荧光技术研究了KDTN对HSA构象的影响,结果表明,KDTN的加入对HSA构象影响不大。     

Characterization of the interaction between farrerol and bovine serum albumin by fluorescence and circular dichroism

The binding of farrerol to bovine serum albumin (BSA) in aqueous solution was investigated by fluorescence quenching spectra, synchronous fluorescence spectra, circular dichroism (CD) and the three-dimensional (3D) fluorescence spectra at pH 7.40. The results of fluorescence titration indicated that farrerol could quench the intrinsic fluorescence of BSA in a static quenching way. The cause of showing upward curvy patterns in Stern-Volmer plots was analyzed. The binding sites number n and binding constant K using fluorescence quenching equation at 310 K were calculated. The binding distance and the energy transfer efficiency between farrerol and BSA were also obtained according to the theory of F?rster's non-radiation energy transfer. The effect of some metal ions on the binding constant of farrerol with BSA was also studied. The effect of farrerol on the conformation of BSA was analyzed using CD, synchronous fluorescence spectra and three-dimensional (3D) fluorescence spectra under experimental conditions. Furthermore, the fluorescence displacement experiments indicated that farrerol could bind to the site I of BSA.     

分子光谱法研究铝酞菁与牛血红蛋白的相互作用

利用紫外可见吸收光谱和荧光光谱研究了在生理pH条件下铝酞菁与牛血红蛋白(BHb)的相互作用.实验结果表明:铝酞菁分子与BHb发生反应生成基态复合物,导致BHb内源荧光的猝灭,该猝灭属于静态猝灭.测定了不同温度下该反应的表观结合常数、结合位点数及结合热力学参数,热力学参数的变化表明铝酞菁与BHb之间以静电和疏水作用力为主;根据Frster能量转移理论,测得供体与受体间结合距离r和能量转移效率E;并用同步荧光光谱法探讨了铝酞菁对BHb构象的影响.     

Investigation of the association behaviors between biliverdin and bovine serum albumin by fluorescence spectroscopy

The interaction between biliverdin and bovine serum albumin (BSA) has been studied by steady fluorescence spectroscopy, synchronous fluorescence and resonance light scanning spectra. The binding of biliverdin to BSA quenches the tryptophan residue fluorescence and the results show that both static and dynamic quenching occur together with complex formation. The binding constant and binding sites of biliverdin to BSA at pH 7.1 are calculated to be 3.33 × 10⁸ L/mol and 1.54, respectively, according to the double logarithm regression curve. In addition, the distance between the biliverdin and BSA is estimated to be 1.25 nm using Föster's equation on the basis of the fluorescence energy transfer. Furthermore the synchronous fluorescence spectra show that the microenvironment of the tryptophan residues has not obvious changes, which obeys the phase distribution model. Finally, the thermodynamic data show that biliverdin molecules enter the hydrophobic cavity of BSA via hydrophobic interaction.     

荧光光谱法研究青蒿素类抗疟药与CYP2B6酶的相互作用

在模拟人体生理条件下,采用荧光光谱法及同步荧光法研究了抗疟药青蒿素(ART)和双氢青蒿素(DHA)与药物代谢酶CYP2B6之间的相互作用.结果表明,在温度为296～303 K时,ART和DHA对CYP2B6的猝灭机制均以静态猝灭为主,当温度由303 K升高至310 K时,则为静态猝灭和动态猝灭共同作用的混合猝灭机制;在...     

阿霉素与牛血清白蛋白结合作用的研究

结合荧光光谱和吸收光谱研究了阿霉素与牛血清白蛋白间的结合作用，确定了 阿霉素对牛血清白蛋白的荧光猝灭过程的猝灭机理，测定了不同酸度条件下该结合 反应的结合常数、结合位点数，依据能量转移理认确定了药物和蛋白间的结合距离 。通过比较阿霉素和去糖基阿霉素与牛血清白蛋白的相互作用，结合阿霉素对蛋白 构象的影响，讨论了药物与蛋白的结合模式。