稀土荧光共聚物与蛋白质作用机理的光谱研究

用含铕稀土配合物单体成功地合成出含铕稀土荧光聚合物(PPNEu)。通过荧光光谱仪、紫外可见分光光度计研究了含铕稀土荧光聚合物与牛血清蛋白(BSA)的相互作用机理。结果表明:含铕稀土荧光聚合物能够与BSA相互结合,并使BSA出现荧光猝灭现象,该猝灭效应属于静态猝灭;结合常数和结合位点数分别为:KA=8.49×104 L/mol,n=1.25(298K);KA=7.45×104 L/mol,n=1.25(308K);在与BSA的结合过程中,吉布斯自由能(ΔG)为负值,熵变(ΔS)为正值且焓变(ΔH)为负值,说明静电力在PPNEu与BSA结合过程中起着主要作用。基于Frster理论,结合距离为3.38nm。     

Pd(Ⅱ)与牛血清白蛋白相互作用的荧光光谱研究

采用荧光光谱法研究了Pd(Ⅱ)与牛血清白蛋白(BSA)的相互作用.使用高斯多峰拟合法对Pd(Ⅱ)-BSA体系荧光光谱的各荧光成分进行了解析.结果表明,BSA的荧光主要来源于色氨酸(Trp)残基,并推测Pd(Ⅱ)与BSA结合作用的位置在第212位Trp残基上.研究了Pd(Ⅱ)与BSA相互作用的荧光猝灭光谱,通过紫外吸收光谱的变化和猝灭速率常数Kq的比较对Pd(Ⅱ)与BSA相互作用的荧光猝灭机理进行了判别.结果表明,Pd(Ⅱ)对BSA的荧光猝灭属于静态猝灭.     

采用荧光光谱法研究DDAF与BSA的相互作用

研究新型反离子季铵盐DDAF与BSA的相互作用。通过单因素法,选出适宜的作用时间、Na+浓度和pH值;并采用荧光光谱法和三维荧光光谱法研究DDAF与BSA的相互作用机制。作用时间15 min、钠离子浓度为0.04 mol/L、pH值为待测液的初始pH值为适宜的实验条件;不同温度(290.15,296.15,303.15,310.15 K)下二者的双分子猝灭常数(Kq)分别为7.03×10~(10),8.53×10~(10),1.09×10~(11),1.12×10~(11) L/(mol·s);结合位点数(n)分别为1.17、0.84、0.97和1.02;?H和?S均大于0;三维荧光光谱显示,加入DDAF后,BSA的荧光峰发生蓝移,且荧光强度下降16.63%。DDAF和BSA为混合猝灭;二者以1∶1结合;二者间的作用力是疏水作用力;DDAF使BSA氨基酸残基周围的疏水环境增强。     

人血清蛋白与重金属离子相互作用的荧光光谱

通过荧光光谱技术研究重金属离子与人血清蛋白(HSA)间的结合作用机制.测量了人血清蛋白与重金属离子Pb2+,Cr6+,Cu2+在290K和300K温度下相互作用的荧光光谱,建立猝灭方程.3种重金属离子与HSA的猝灭均属于静态猝灭,根据静态猝灭方程Stern-Volmer分别计算出290K和300K温度下HSA与3种重金属离子相互作用结合常数,290K下结合常数Ksv分别为1.731×103,5.580×103,4.461×104;300K下结合常数Ksv分别为1.354×103,5.418×103,4.461×104,结合位点数分别为1.237,1.528,0.506.证明了重金属离子Pb2+,Cr6+,Cu2+与HSA之间的相互作用是自发的.     

三维荧光光谱法和圆二色谱法研究氨基比林与牛血清白蛋白分子的相互作用

该文在模拟生理条件下,采用荧光光谱法、三维荧光光谱法以及圆二色谱法,研究氨基比林(PYM)与牛血清白蛋白(BSA)之间的相互作用。研究结果表明PYM对BSA有强烈的荧光猝灭作用,其荧光猝灭机制为静态猝灭。由热力学参数判定PYM和BSA的主要作用力为氢键和范德华力,并且相互作用是自发进行的。根据F觟rster的偶极-偶极非辐射能量转移理论可以得出PYM与BSA之间的结合距离为2.09 nm。利用三维荧光光谱和圆二色谱技术,分析PYM对BSA蛋白构象的影响,表明PYM与BSA相互作用后使BSA的微环境和构象发生改变。     

分子量为3000的壳聚糖与牛血清白蛋白相互作用的研究

在模拟人体生理条件下,利用荧光光谱、圆二色谱、紫外-可见吸收光谱法和三维荧光法研究了3 000分子量的壳聚糖(CS)与牛血清白蛋白(BSA)的相互作用。结果表明,CS对BSA的紫外吸收光谱具有增强作用,而对荧光光谱具有较强的荧光猝灭作用且峰位明显蓝移8~10 nm。用Stern-Volmer方程分别对实验数据进行分析,得出结论,CS对BSA的荧光猝灭作用是属于静态荧光猝灭;与其反应生成了新的复合物,发生了分子内的非辐射能量转移;并求得相互作用过程的结合常数KA(Kb)和热力学参数(ΔG、ΔH、ΔS),确定了它们之间的主要作用力是静电作用力,但疏水作用也不可忽略。圆二色谱、同步荧光光谱和三维荧光光谱法表明了CS对牛血清白蛋白的构象和所处的微环境发生了一定程度的变化。     

多光谱法探究壳聚糖与酪蛋白的结合作用

目的 以壳聚糖(Chitosan,CS)和酪蛋白(Casein,CA)为研究对象,通过探究多糖对蛋白质结构的影响机制,以期改进多糖对蛋白质食品贮藏的效果。方法 采用荧光光谱法和傅里叶变换红外光谱法,在不同pH条件下,研究CS与CA的结合反应,通过数学方程计算其结合常数、结合位点数和结合作用力。结果 不同pH值条件下,CS对CA均产生荧光猝灭,均属于静态猝灭,pH=5.2时猝灭常数达到最大值4.702×10³ L/mol,且结合常数达到最大值9.914×10³ L/mol。热力学分析表明,结合反应自发进行属于放热反应,主要结合驱动力为静电相互作用。同步荧光光谱和三维荧光光谱表明,CS与CA的相互作用能够使CA的空间构象发生改变。傅里叶变换红外光谱说明CS加入和pH值的变化导致了CA二级结构发生变化。结论 CS与CA结合会导致CA的结构发生变化,这为蛋白质–多糖相互作用提供一定的理论依据,也为以蛋白质–多糖为基础的乳制品贮藏及开发提供实验依据。     

荧光光谱及分子模拟研究β-榄香烯与牛血清白蛋白的相互作用

在模拟生理p H条件(p H=7.40)下,用荧光光谱和分子模拟法研究β-榄香烯与牛血清白蛋白(BSA)的相互作用。在308 K和318 K温度下,激发波长(λex)为280 nm,测定BSA在340 nm的内源性荧光强度随着β-榄香烯浓度增加的变化,用分子对接方法研究β-榄香烯与牛血清白蛋白(BSA)的相互作用。β-榄香烯与牛血清白蛋白的反应机制为静态猝灭,作用力类型为疏水作用。分子模拟结果表明:β-榄香烯与牛血清白蛋白亚结构域A结合,二者之间有疏水作用和静电作用,且以疏水作用为主,这与荧光光谱结果一致。β-榄香烯与BSA具有较强的相互作用,以血清白蛋白为载体,β-榄香烯作为药物可通过血液循环到达病变部位,发挥药效。     

碳量子点的制备及与牛血清蛋白的相互作用

以木炭为碳源,分别采用回流、微波及超声等不同方法制备碳量子点(Carbon quantum dots,CQDs)。比较不同方法的优劣并优化反应条件、考察不同因素对其荧光量子产率的影响。得到最佳制备方法,制得粒径较小的荧光CQDs,用钝化剂PEG2000修饰后,提高其荧光寿命和量子产率。将修饰后的CQDs应用于与牛血清白蛋白(Bovine serum albumin,BSA)的相互作用,采用紫外吸收光谱法和荧光光谱法探讨其相互作用机理。结果表明,经回流法所制CQDs的荧光量子产率最高,其与BSA之间的荧光猝灭为静态猝灭过程。     

荧光光谱法研究头孢孟多脂与牛血清白蛋白的相互作用

模拟生理条件下,用荧光光谱法和紫外-可见吸收光谱法研究头孢孟多酯和牛血清白蛋白(BSA)结合反应的特征。研究表明:头孢孟多酯与BSA形成复合物,从而猝灭BSA的内源性荧光,该过程为静态猝灭过程。根据SternVolmer方程得出不同温度下结合位点数n和结合常数Ka;结合位点位于BSA的亚结构IIA中。通过计算相应的热力学参数,确定头孢孟多酯与BSA之间的作用力主要为静电作用力。利用同步荧光光谱探讨了头孢孟多酯与BSA作用前后白蛋白的构型变化。Hill系数nH1,表明头孢孟多酯有弱的负协同作用。此研究不仅对于揭示体内药物动力学问题和指导临床合理用药具有一定意义,而且对药物分子设计及新药开发等也具有重要指导意义。     

Interaction of malachite green with bovine serum albumin: determination of the binding mechanism and binding site by spectroscopic methods

The interaction between malachite green (MG) and bovine serum albumin (BSA) under simulative physiological conditions was investigated by the methods of fluorescence spectroscopy, UV-vis absorption and circular dichroism (CD) spectroscopy. Fluorescence data showed that the fluorescence quenching of BSA by MG was the result of the formation of the MG-BSA complex. According to the modified Stern-Volmer equation, the effective quenching constants (K(a)) between MG and BSA at four different temperatures were obtained to be 3.734 x 10(4), 3.264 x 10(4), 2.718 x 10(4), and 2.164 x 10(4)L mol(-1), respectively. The enthalpy change (Delta H) and entropy change (DeltaS) were calculated to be -27.25 kJ mol(-1) and -11.23 J mol(-1)K(-1), indicating that van der Waals force and hydrogen bonds were the dominant intermolecular force in stabilizing the complex. Site marker competitive experiments indicated that the binding of MG to BSA primarily took place in sub-domain IIA. The binding distance (r) between MG and the tryptophan residue of BSA was obtained to be 4.79 nm according to F?rster theory of non-radioactive energy transfer. The conformational investigation showed that the presence of MG decreased the alpha-helical content of BSA (from 62.6% to 55.6%) and induced the slight unfolding of the polypeptides of protein, which confirmed some micro-environmental and conformational changes of BSA molecules.     

Spectroscopic and electrochemical studies on the molecular interaction between copper sulphide nanoparticles and bovine serum albumin

The interaction of covellite hexagonal phase of copper sulphide nanoparticles (CuS NPs) with bovine serum albumin (BSA) was examined systematically by using fluorescence, UV–visible, circular dichroism (CD), Fourier transform infrared (FTIR), dynamic light scattering (DLS) and molecular modelling techniques. Electrochemical method was studied to further confirm the interaction of BSA with CuS NPs. The results of fluorescence studies demonstrated that fluorescence of BSA was quenched by CuS NPs via a static quenching mechanism. The negative values of thermodynamic parameters (ΔG, ΔH and ΔS) indicated that the binding process is spontaneous, exothermic and van der Waals force or hydrogen bonding plays major roles in the interaction of CuS NPs with BSA. The interaction of CuS NPs with Trp residue was established by synchronous studies, and competitive binding studies revealed that Trp-212 of subdomain IIA was involved in the interaction with these nanoparticles. Further, the efficiency of energy transferred and the distance between fluorophore (BSA) and acceptor (CuS NPs) were calculated using Forster’s resonance energy transfer theory. The results of UV–visible, CD, FTIR and DLS revealed that the CuS NPs interact with BSA by inducing the conformational changes in secondary structure and reducing the α-helix content of BSA. Molecular modelling studies suggested that CuS NPs bind to site I of sub domain IIA of BSA. The results of spectroscopic and molecular docking studies were complimented by the electrochemical techniques.     

Temperature-swing adsorption of proteins in water using N-isopropylacrylamide copolymer gel particles

The adsorption and desorption behaviors of bovine serum albumin (BSA) in water for temperature-responsive polymer gel particles have been investigated by the temperature-swing operation between 298 and 313 K, where the cationic N-isopropylacrylamide (NIPA) gels copolymerized with vinylbenzyl trimethylammonium chloride (VBTA) or 2-(dimethylamino)ethyl methacrylate (DMAEMA) were used. The NIPA-VBTA and the NIPA-DMAEMA copolymer gels adsorbed BSA while the NIPA homopolymer gel hardly adsorbed BSA, indicating that the copolymer gels adsorb BSA through the electrostatic attraction between the positively charged groups in the gels and the negatively charged BSA. The adsorption amounts for the NIPA-DMAEMA gels were smaller than those for the NIPA-VBTA gels. This may be because almost every VBTA group, which is a quaternary ammonium salt, can be positively charged in water, while only some of the tertiary amine DMAEMA groups are protonated in water. Moreover, it was found that both the copolymer gels with a large mesh size of the polymer network repeatedly adsorbed BSA at 298 K and desorbed some of pre-adsorbed BSA at 313 K by the temperature-swing operation. This BSA desorption may result from the decrease of the number of the positively charged groups accessible to BSA due to the shrinking of the constituent polymer chains.     

The interaction between 4-aminoantipyrine and bovine serum albumin: multiple spectroscopic and molecular docking investigations

4-Aminoantipyrine (AAP) is widely used in the pharmaceutical industry, in biochemical experiments and in environmental monitoring. AAP as an aromatic pollutant in the environment poses a great threat to human health. To evaluate the toxicity of AAP at the protein level, the effects of AAP on bovine serum albumin (BSA) were investigated by multiple spectroscopic techniques and molecular modeling. After the inner filter effect was eliminated, the experimental results showed that AAP effectively quenched the intrinsic fluorescence of BSA via static quenching. The number of binding sites, the binding constant, the thermodynamic parameters and binding subdomain were measured, and indicated that AAP could spontaneously bind with BSA on subdomain IIIA through electrostatic forces. Molecular docking results revealed that AAP interacted with the Glu 488 and Glu 502 residues of BSA. Furthermore, the conformation of BSA was demonstrably changed in the presence of AAP. The skeletal structure of BSA loosened, exposing internal hydrophobic aromatic ring amino acids and peptide strands to the solution.     

Interactions of vitamin K3 with herring-sperm DNA using spectroscopy and electrochemistry

By means of ultraviolet-visible (UV-Vis) and fluorescence spectra, the binding ratio between vitamin K(3) and herring-sperm DNA in a physiological pH environment (pH = 7.40) was determined as n(K3):n(DNA) = 2:1, and the binding constants of vitamin K(3) binding to DNA at different temperatures were determined as K(θ)(298K) = 1.28 × 10(5) L·mol(-1) and K(θ)(310K) = 7.19 × 10(4) L·mol(-1), which were confirmed using the double reciprocal method are Δ(r)H(m)(θ) = -3.57 × 10(4) J·mol(-1), Δ(r)G(m)(θ) = -2.92 × 10(4) J·mol(-1), and Δ(r)S(m)(θ) = 217.67 J·mol(-1)K(-1). The driving power of this process was enthalpy. An intercalation binding of the vitamin K(3) with DNA was supported by a competitive experiment using acridine orange (AO) as a spectral probe. By combination analysis of the Scatchard method and cyclic voltammetry, we suggested that the interaction mode between vitamin K(3) and herring-sperm DNA would be a mixed mode. The quinonoid, duality fused-ring of vitamin K(3) can intercalate into the base pairs of DNA, and there is an electrostatic binding along with intercalation binding.     

Studies on binding of single‐stranded DNA with reduced graphene oxide–silver nanocomposites

The binding reaction of reduced graphene oxide–silver nanocomposites (rGO–AgNCs) with calf thymus single‐stranded DNA (ssDNA) was studied by ultraviolet–visible absorption, fluorescence spectroscopy and circular dichroism (CD), using berberine hemisulphate (BR) dye as a fluorescence probe. The absorbance of ssDNA increases, but the fluorescence intensity is quenched with the addition of rGO–AgNCs. The binding of rGO–AgNCs with ssDNA was able to increase the quenching effects of BR and ssDNA, and induce the changes in CD spectra. All of the evidence indicated that there was a relatively strong interaction between ssDNA and rGO–AgNCs. The data obtained from fluorescence experiments revealed that the quenching process of ssDNA caused by rGO–AgNCs is primarily due to complex formation, i.e. static quenching. The increasing trend of the binding equilibrium constant (K a) with rising temperature indicated that the binding process was an endothermic reaction. The calculated thermodynamic parameters showed that the binding process was thermodynamically spontaneous, and hydrophobic association played predominant roles in the binding of ssDNA to the surface of rGO–AgNCs.Inspec keywords: DNA, fluorescence spectroscopy, circular dichroism, nanofabrication, nanobiotechnology, silver, hydrophobicity, molecular biophysics, radiation quenching, biochemistry, nanocomposites, visible spectra, ultraviolet spectra, dyes, fluorescence, association, graphene compoundsOther keywords: ssDNA increases, fluorescence intensity, fluorescence experiments, binding equilibrium constant, binding process, reduced graphene oxide‐silver nanocomposites, binding reaction, calf thymus single‐stranded DNA, fluorescence spectroscopy, fluorescence probe, complex formation, static quenching, calculated thermodynamic parameters, rGO‐AgNCs binding, ultraviolet‐visible absorption, berberine hemisulphate dye, circular dichroism, CO‐Ag     

荧光光谱法研究桑色素与人血清白蛋白的相互作用

采用荧光猝灭光谱、紫外-可见吸收光谱研究了桑色素与人血清白蛋白(HSA)的结合作用。实验表明桑色素对HSA的荧光猝灭属于单一静态猝灭反应,在溶液中以摩尔比1:1牢固结合,各结合反应的平衡常数Kp>105,结合常数Kb>104;根据F rster非辐射能量转移机理,求算HSA与桑色素间距离r为3.81~3.58nm,能量转移效率E为0.18~0.13。并根据结合反应的热力学常数推测了药物与HSA之间的主要作用力类型为疏水作用力和偶极-偶极作用力。     

Study on protein conformation and adsorption behaviors in nanodiamond particle-protein complexes

In the present research, the conformation of bovine serum albumin (BSA) in the nanodiamond particle (ND)-BSA complex was studied by Fourier transform infrared spectroscopy, fluorescence spectroscopy, UV-vis spectroscopy, and circular dichroism spectroscopy. The spectroscopic study revealed that most BSA structural features could be preserved in the complex though the BSA underwent conformational changes in the complex due to ND-BSA interaction. In addition, BSA adsorption isotherms and zeta-potential measurements were employed to investigate the pH dependence of the ND-BSA interaction. The changes in surface charge of the ND-BSA complex with pH variations indicated that the binding of BSA to ND might lead to not only the adsorption of BSA onto the ND surface but also the partial breakup of ND aggregates into relatively small ND-BSA aggregates because of the strong binding force between ND and BSA. The results show that ND is an excellent platform for protein immobilization with high affinity and holds great potential to be used for biosensor applications.