Investigation on the interaction of glipizide with bovine hemoglobin by spectroscopy and molecular docking

This study aims to investigate the interaction between glipizide and bovine hemoglobin using fluorescence quenching, circular dichroism spectroscopy in various temperatures (293, 303, and 310?K) and molecular docking methods. The results demonstrated that glipizide could cause strong fluorescence quenching of bovine hemoglobin by a dynamic quenching mechanism, during which the hydrophobic interaction played a dominant role in this system. The order of magnitude of binding constant is 10⁴, and the number of binding site in the system was close to 1. It also showed that tyrosine residues and tryptophan residues were both involved in the binding of glipizide with bovine hemoglobin, and was closer to the later. Circular dichroism spectra revealed that the conformation of bovine hemoglobin was changed during the binding reaction. The interaction of the system was studied by both spectroscopic method and molecular docking simulation, and the conclusions are consistent.     

Toxic effects of copper (II) ions on bovine hemoglobin

In this paper, the toxic influence of copper ions (II) on bovine hemoglobin was investigated by the combination of ultraviolet-visible absorption, fluorescence, time-resolved fluorescence, synchronous fluorescence, and circular dichroism spectra. Driven by hydrophobic and electrostatic forces, copper ions (II) could interact with bovine hemoglobin to form bovine hemoglobin-copper ions (II) complex with one binding site. The binding constant (K) was 1.57?×?10⁴, 1.89?×?10⁴ and 2.11?×?10⁴?L/mol at 298, 304, and 310?K, respectively. The binding distance (r) was 4.24?nm. Fluorescence and time-resolved fluorescence spectra showed that bovine hemoglobin quenched by copper ions (II) was a static quenching process. Results of synchronous fluorescence spectra revealed that the microenvironment and the conformation of bovine hemoglobin were changed during the binding reaction. Data of circular dichroism spectra suggested that with the increasing concentration of copper ions (II), the secondary structure of bovine hemoglobin underwent a decrease in α-helix and alteration in backbone microenvironment. Copper ions (II) was thus evidenced to have a certain toxic effect on physical bodies.     

Spectroscopic Studies on the Thermodynamics of L-Cysteine Capped CdSe/CdS Quantum Dots��BSA Interactions

CdSe/CdS quantum dots (QDs) capped with L-cysteine can provide an effective platform for the interactions with bovine serum albumin (BSA). In this study, absorption and fluorescence (FL) spectroscopy were used to study the binding reactions of QDs with BSA, respectively. The binding constant (??10⁴ M^-1) from FL quenching method matches well with that determined from the absorption spectral changes. The modified Stern-Volmer quenching constant (5.23?×?10⁴, 5.22?×?10⁴, and 4.90?×?10⁴ M^-1) and the binding sites (??1) at different temperatures (304 K, 309 K, and 314 K) and corresponding thermodynamic parameters were calculated (?G?<?0, ?H?<?0, and ?S?<?0). The results show the quenching constant is inversely correlated with temperature. It indicates the quenching mechanism is the static quenching in nature rather than dynamic quenching. The negative values of free energy (?G?<?0) suggest that the binding process is spontaneous, ?H?<?0 and ?S?<?0 suggest that the binding of QDs to BSA is enthalpy-driven. The enthalpy and entropy changes for the formation of ground state complex depend on the capping agent of QDs and the protein types. Furthermore, the reaction forces were discussed between QDs and BSA, and the results show hydrogen bonds and van der Waals interactions play a major role in the binding reaction.     

Investigation of the Interaction Between Rutin and Trypsin in Solution by Multi-Spectroscopic Method

ABSTRACT

The interactions between rutin and trypsin were investigated by UV-Vis absorption, CD, fluorescence, resonance light-scattering spectra, synchronous fluorescence, and three-dimensional fluorescence spectra techniques under physiological pH 7.40. Rutin effectively quenched the intrinsic fluorescence of trypsin via static quenching. The enthalpy change and entropy change were estimated to be ?8.23 kJ·mol^?1 and 53.66 J·mol^?1·K^?1 according to the van't Hoff equation. The process of binding rutin to trypsin was a spontaneous molecular interaction procedure. This result indicates that hydrophobic and electrostatic interactions played a major role in stabilizing the complex. The conformation of trypsin was discussed by CD, synchronous, and three-dimensional fluorescence techniques.     

Interaction of Sulfadiazine with Model Water Soluble Proteins: A Combined Fluorescence Spectroscopic and Molecular Modeling Approach

The binding behavior of antibacterial drug sulfadiazine (SDZ) with water soluble globular proteins like bovine as well as human serum albumin (BSA and HSA, respectively) and lysozyme (LYS) was monitored by fluorescence titration and molecular docking calculations. The experimental data reveal that the quenching of the intrinsic protein fluorescence in presence of SDZ is due to the strong interaction in the drug binding site of the respective proteins. The Stern-Volmer plot shows positive deviation at higher quencher concentration for all the proteins and was explained in terms of a sphere of action model. The calculated fluorophore-quencher distances vary within 4?~?11 Å in different cases. Fluorescence experiments at different temperature indicate thermodynamically favorable binding of SDZ with the proteins with apparently strong association constant (~10⁴–10⁵ M^?1) and negative free energy of interaction within the range of ?26.0?~??36.8 kJ mol^?1. The experimental findings are in good agreement with the respective parameters obtained from best energy ranked molecular docking calculation results of SDZ with all the three proteins.     

Synthesis of Amide Compounds of Ferulic Acid and Their Association with Bovine Serum Albumin

ABSTRACT

In this work, three new amide compounds of ferulic acid (FA) were synthesized. The fluorescence and ultraviolet spectroscopy were explored to study the interactions between three amide compounds of FA and bovine serum albumin (BSA) under imitated physiological conditions. The experimental results showed that the fluorescence quenching mechanism between BSA and three amide compounds of FA were mainly static quenching and nonradiation energy transfer at 25°C, 30°C, and 37°C. The Stern–Volmer quenching constants, the binding constants, and the number of binding sites and corresponding thermodynamic parameters ΔH, ΔG, and ΔS were calculated at different temperatures. From the thermodynamic parameters, we concluded that the action force was mainly a hydrophobic interaction. According to the F?rster theory of nonradiation energy transfer, the binding distances (r) between BSA and amide compounds are less than 7 nm. Furthermore, the effects of amide compounds on the conformation of BSA were analyzed using synchronous fluorescence spectroscopy.     

Mechanism of the toxicological interactions of decabrominated diphenyl ether with hemoglobin

Polybrominated diphenyl ethers are vital flame retardants in production and human life and they are widespread global organic pollutants in the environment and likewise in food and feed, causing a potential health concern. Hemoglobin is the main protein in the blood, which is a carrier of oxygen in red blood cells. This work aimed at investigating the toxic interactions of polybrominated diphenyl ethers with hemoglobin, by molecular modeling and spectroscopic analysis methods. Decabrominated diphenyl ether and bovine hemoglobin were selected as representatives for polybrominated diphenyl ethers and hemoglobin, respectively. The experimental results indicated that decabrominated diphenyl ether changed the frames’ conformation and the microenvironment of bovine hemoglobin, which can affect the physiological function of the protein. Decabrominated diphenyl ether combined with bovine hemoglobin with the average number of binding sites (0.7) to form bovine hemoglobin–decabrominated diphenyl ether complex. The binding constant was 860.95?L mol^?1. In addition, the molecular docking data revealed that the van der Waals forces played the primary role in the interaction between bovine hemoglobin and decabrominated diphenyl ether. The study provides insight into the molecular toxicity mechanism of decabrominated diphenyl ether during the blood transportation in vivo.     

Interaction Between Isoquercitrin and Bovine Serum Albumin by a Multispectroscopic Method

ABSTRACT

The interaction of isoquercitrin and bovine serum albumin (BSA) was investigated by means of fluorescence spectroscopy (FS), resonance light scattering spectroscopy (RLS), and ultraviolet spectroscopy (UV). The apparent binding constants (K _a) between isoquercitrin and BSA were 5.37 × 10⁵ L mol^?1 (293.15 K) and 2.34 × 10⁵ L mol^?1 (303.15 K), and the binding site values (n) were 1.18 ± 0.03. According to the Förster theory of non-radiation energy transfer, the binding distances (r) between isoquercitrin and BSA were 1.94 and 1.95 nm at 293.15 K and 303.15 K, respectively. The experimental results showed that the isoquercitrin could be inserted into the BSA, quenching the inner fluorescence by forming the isoquercitrin–BSA complex. The addition of increasing isoquercitrin to BSA solution leads to the gradual enhancement in RLS intensity, exhibiting the formation of the aggregate in solution. It was found that both static quenching and non-radiation energy transfer were the main reasons for the fluorescence quenching. The entropy change and enthalpy change were negative, which indicated that the interaction of isoquercitrin and BSA was driven mainly by van der Waals interactions and hydrogen bonds. The process of binding was a spontaneous process in which Gibbs free energy change was negative.     

Study on the Interaction Between Water-Soluble CdTe Quantum Dots and Ovalbumin by Optical Spectroscopy

ABSTRACT

CdTe quantum dots (QDs) modified by 2-mercaptoethylamine hydrochloride (CA) and thioglycolic acid (TGA), respectively, were synthesized in aqueous medium. The interaction of CdTe QDs with ovalbumin has been investigated in depth by Fourier transform infrared spectroscopy (FTIR), UV-Vis absorption, fluorescence-quenching spectrometry, and resonance Rayleigh scattering spectroscopy (RRS). Fluorescence data show that the quenching type of ovalbumin by CA-CdTe QDs is static quenching with the binding constant being 10^?4 M^?1, and the number of binding sites being one. The calculated thermodynamic parameters demonstrate that the main binding forces are hydrophobic interaction and electrostatic attraction. In contrast, the quenching style of ovalbumin by TGA-CdTe QDs is verified to be dynamic quenching. Under suitable acidity conditions, the interaction of CA-CdTe QDs or TGA-CdTe QDs with ovalbumin leads to the remarkable enhancement of RRS, and the increments are found to be proportional to the concentration of ovalbumin in a certain range. A simple and highly sensitive RRS approach for determining ovalbumin is developed. In addition, the causes for the enhancement of RRS and the quenching of fluorescence are investigated to shed light on the quenching mechanism.     

Study on the Interaction between Florasulam and Bovine Serum Albumin

In this paper, the interaction between florasulam (FU, 2′,6′,8-trifluoro-5-methoxy [Kragh-Hansen U, Molecular aspects of ligand binding to serum albumin. Pharmacol Rev 33(1):17–53 1981; Carter DC and Ho JX, Structure of serum albumin. Adv Protein Chem 45:153–203 1994; He XM, and Carter DC, Atomic structure and chemistry of human serum albumin. Nature 358(6383):209–215 1992] triazolo [1,5-c]pyrimidine-2-sulfonanilide) and bovine serum albumin (BSA) was investigated by fluorescence, ultraviolet absorption (UV) and Far-UV circular dichroism (CD) spectrometries. A strong fluorescence quenching was observed and the quenching mechanism was considered as static quenching. The binding constant of FU with BSA at 299 and 309 K were obtained as 1.5?×?10⁴ and 7.1?×?10³ l mol^?1, respectively. There was one binding site between FU and BSA. The thermodynamic parameters enthalpy change (ΔH) and entropy change (ΔS) were calculated as ?57.89 kJ mol^?1 and ?113.6 J mol^?1 K^?1, respectively, which indicated that the acting force between FU and BSA was mainly hydrogen bond and Van der Waals force. According to the Förster non-radiation energy transfer theory, the average binding distance between donor (BSA) and acceptor (FU) was obtained (r?=?1.59 nm). The investigations of the UV/Vis and CD spectra of the system showed that the conformation of BSA was changed in presence of FU.     

Interaction between flavin mononucleotide-containing azoreductase and azo dyes

ABSTRACT

Azoreductase, a flavin mononucleotide-containing oxidoreductase from Escherichia coli, can catalyze the reduction of azo dyes to form aromatic amine compounds. Few spectroscopic studies have explored the binding mode of azo dyes or the role of the arginine at site 59 in Azoreductase. In this article, protein engineering strategy has been used to construct one mutant in which the arginine residue at site 59 was mutated to glycine. Fluorescence spectroscopic data showed that the addition of Methyl Red and Methyl Orange resulted in the fluorescence quenching of the cofactor flavin mononucleotide bound to Azoreductase. The association constant was fitted using the standard binding equation instead of the Stern-Volmer equation. The results showed that the mutation from the arginine to glycine at site 59 weakened the association constant from 2.21?×?10⁵?L.mol^?1 to 4.55?×?10⁴?L.mol^?1 at 25°C. A similar phenomenon was also observed when Methyl Orange was used as a substrate. In each case, the association constant tended to decrease as the temperature increased from 25°C to 37°C. Thermodynamic parameter analysis revealed that the interaction type changed from a van der Waals interaction (between Azoreductase and the dyes) to a hydrogen bonding interaction (between the mutant and the dyes). Moleculcar docking was also performed in this work to provide some support for the binding mode and binding stability between Azoreductase/mutant and azo dyes.     

Quenching of porous silicon photoluminescence by molecular oxygen and dependence of this phenomenon on storing media and method of preparation of pSi photosensitizer

The quenching of porous silicon photoluminescence (pSi PL) by molecular oxygen has been studied in different storing media in an attempt to clarify the mechanism of the energy transfer from the silicon photosensitizer to the oxygen acceptor. Luminescent materials have been prepared by two methods: electrochemical anodizing and chemical etching. Different structural forms were used: porous layers on silicon wafer and two kinds of differently prepared powder. Dry air and liquid water were employed as storing media; quenching behaviour was under observation until total degradation of quenching properties. Singlet oxygen molecules generation through energy transfer from photoluminescent pSi was the only photosensitizing mechanism observed under dry gas conditions. This PL quenching process was preferentially developed at 760 nm (1.63 eV) that corresponds to the formation of the ¹Σ singlet oxygen state. Oxidation of the pSi photosensitizer was the main factor that led to its total deactivation in a time scale of few weeks. Regarding water medium, different photosensitizing behaviour was observed. In watery conditions, two preferred energy levels were found: the one detected in dry gas and another centred at approximately 2.2 eV (550 nm). Formation of reactive oxygen species (ROS) different from singlet oxygen, such as superoxide anion or superoxide radical, can be responsible for the second one. This second quenching process developed gradually after the initial contact of pSi photosensitizer with water and then degraded. The process lasted only several hours. Therefore, functionalization of the pSi photosensitizer is probably required to stabilize its PL and quenching properties in the watery physiological conditions required for biomedical applications.     

A combined experimental and theoretical study of the pH‐dependent binding mode of NAD+ by water‐soluble molecular clips

The highly selective recognition process of NAD⁺ and NADH (as important cofactors of many redox enzymes) by molecular clips in aqueous solution is studied systematically by a combined experimental and quantum‐chemical approach. The strongly pH‐dependent complexation‐induced ¹H NMR shifts of the guest molecule indicate that in buffered aqueous solution at pH = 7.2 the nicotinamide ring, the active site of NAD⁺, is preferentially bound inside the cavity of the molecular clip, whereas in pure water under slightly acidic conditions both units (the nicotinamide ring as well as the adenine moiety) are located outside the cavity. The latter finding is explained by a competing self‐aggregation of NAD⁺ which prohibits the recognition process. In addition, the investigation of the NAD⁺ fragments NMNA, NMN, and AMP as well as the comparison of measured and computed chemical shieldings provides information on possible binding modes. Under equal conditions the binding of NADH to the molecular clip is significantly weaker than that of NAD⁺, so that the oxidation states (NAD⁺/NADH) can be distinguished by the molecular clips. The nucleotides NMN and AMP are bound less strongly by the molecular clips than NAD⁺. The weaker binding indicates that multiple aromatic π?π and cation?π host–guest interactions only possible in NAD⁺ have a synergetic effect on the complex stability. In addition to the inhibition of the cofactor NAD⁺, a further implication is the development of sensors since a quenching of fluorescence is observed for specific molecular clips by the addition of NAD⁺. Copyright © 2009 John Wiley & Sons, Ltd.     

Influence on drug efficacy of the binding behavior of pioglitazone hydrochloride with tryptophan residues,and tyrosine residues in bovine transferrin

Abstract

The interaction of pioglitazone hydrochloride bound to tryptophan residues and tyrosine residues in bovine transferrin was investigated using synchronous fluorescence spectroscopy at various temperatures (298, 310, and 318?K). From binding constants and thermodynamic parameters, it was shown that 1:1 stable compound was formed by the electrostatic force interaction of pioglitazone hydrochloride bound to tryptophan residues and tyrosine residues in bovine transferrin. The extent of binding between pioglitazone hydrochloride and tryptophan residues in bovine transferrin was more than that between pioglitazone hydrochloride and tyrosine residues in bovine transferrin. At 310?K, the fluorescence quenching ratio number of tyrosine residues and tryptophan residues in bovine transferrin were 47.52% and 54.19%, respectively, which indicated that the fluorescence contribution of tryptophan residues was greater. At 310?K, pioglitazone hydrochloride-tyrosine residues(in bovine transferrin) binding rate were 55.60–73.82%, and the combined model was W?=??0.0315R²???0.1520R?+?0.7385. The value of Hill’s coefficients was greater than 1, which suggested that there was a positive cooperativity between pioglitazone hydrochloride and subsequent ligands. The results of molecular docking were consistent with that of experimental calculation.     

Molecular interaction of silybin with hyaluronidase: A spectroscopic and molecular docking study

In this study, the molecular interaction of silybin with hyaluronidase was investigated by spectroscopic methods and molecular docking. It was found that silybin had strong ability to quench the intrinsic fluorescence of hyaluronidase by a static quenching procedure. The binding constants were obtained at three temperatures (293, 298, and 310 K). The results of synchronous fluorescence and three-dimensional fluorescence and molecular docking showed that silybin bound into the hyaluronidase cavity site and the binding of silybin to hyaluronidase could induce micro-environmental and conformational changes in hyaluronidase, which resulted in the reduced hyaluronidase activity. The thermodynamic parameter analysis and molecular docking experiments revealed that all types of non-covalent interaction, including hydrogen bonding interaction, van der Waals forces, hydrophobic interaction, and electrostatic interaction were present in the binding process of silybin with hyaluronidase. The results obtained here will provide direct evidence at a molecular level to understand the mechanism of the inhibitory effect of silybin against hyaluronidase.     

Interactions between imazethapyr and bovine serum albumin: Spectrofluorimetric study

The interaction between imazethapyr (IMA) and bovine serum albumin (BSA) was investigated by fluorescence spectroscopy. The Stern–Volmer quenching constant (K_SV) at three temperatures was evaluated in order to determine the quenching mechanism. The dependence of fluorescence quenching on viscosity was also evaluated for this purpose. The results showed that IMA quenches the fluorescence intensity of BSA through a static quenching process. The values of the binding constant for the formed BSA–IMA complex and the number of binding sites were found to be 1.51×10⁵ M^?1 and 0.77, respectively, at room temperature. Based on the calculated thermodynamic parameters, the forces that dominate the binding process are hydrogen bonds and van der Waals forces, and the binding process is spontaneous and exothermic. The quenching of protein fluorescence by iodide ion was used to probe the accessibility of tryptophan residues in BSA and the change in accessibility induced by the presence of IMA. According to the obtained results, the BSA–IMA complex is formed in the site where the Trp-134 is located, causing it to become less exposed to the solvent.     

Molecular mechanism investigation on the interaction of Clothianidin with human serum albumin

Abstract

With the widespread application of neonicotinoid insecticides, Clothianidin has received much attention due to the potential harm to human health and ecological environment. However, the mechanism of Clothianidin's underlying toxicity to organisms remains unclear. In this work, the interaction between Clothianidin and human serum albumin was investigated and the intrinsic fluorescence of human serum albumin got quenched via static mechanisms upon the addition of Clothianidin. The binding constants between Clothianidin and human serum albumin at three different temperature were obtained to be 3.543?×?10⁴, 2.995?×?10⁴, and 2.490?×?10⁴ M^?1, respectively. Based on the van't Hoff equation, the thermodynamic parameters, ΔH⁰ and ΔS⁰ were estimated to be ?53.885?KJ mol^?1 and ?110.535?J mol^?1K^?1, respectively. A single binding site was predicted from the binding constants at different temperatures by multiple spectroscopic techniques and the negative values of ΔH⁰ and ΔS⁰ indicated the binding of human serum albumin with Clothianidin was driven by hydrogen bonds or van der Waals forces. Furthermore, the loose and unfolded secondary structure of human serum albumin along with the addition of clothianidin had been observed through ultraviolet-visible absorption and circular dichroism spectra. In addition, it was also found that Clothianidin had polar effects of structural microenviroment not only on Trp but also Tyr residues from synchronous fluorescence analysis. This study illuminates the molecular mechanism of the interaction between human serum albumin and clothianidin for the first time and helps to construct a specific pesticide biosensor system of human health.     

Combined Quenching Mechanism of Anthracene Fluorescence by Cetylpyridinium Chloride in Sodium Dodecyl Sulfate Micelles

The Stern-Volmer quenching constant (K_SV) for quenching of anthracene fluorescence in sodium dodecyl sulfate (SDS) micelles by pyridinium chloride has been reported previously to be 520 M^?1 based on steady state fluorescence measurements. However, such measurements cannot distinguish static versus dynamic contributions to the overall quenching. In the work reported here, the quenching dynamics of anthracene in SDS micelles by cetylpyridinium chloride (CPC), an analogue of pyridinium chloride, were investigated using both steady state and time resolved fluorescence quenching. Concurrent measurement of the decrease in fluorescence intensity and lifetime of anthracene provide a quantitative evaluation of collision induced (i.e. dynamic) versus complex formation (i.e. static) quenching of the anthracene fluorophore. The results reveal that a combined quenching mechanism is operative with approximately equal constants of 249?±?6 M^?1 and 225?±?12 M^?1 for dynamic and static quenching, respectively.     

Biophysical Studies on the Interactions of a Classic Mitochondrial Uncoupler with Bovine Serum Albumin by Spectroscopic,Isothermal Titration Calorimetric and Molecular Modeling Methods

The interaction between a classic uncoupler (2,4-dinitrophenol, DNP) and bovine serum albumin (BSA) was investigated by fluorescence spectroscopy under the physiological conditions. The fluorescence quenching constants were calculated by the Stern-Volmer equation, and based upon the temperature dependence of quenching constants, it was proved that DNP caused a static quenching of the intrinsic fluorescence of BSA. Owing to the static quenching mechanism, different associative binding constants at various temperatures were determined and thus the thermodynamic parameters, namely enthalpy (ΔH = −21.12 kJ mol⁻¹) and entropy changes (ΔS = 23.51 J mol⁻¹ K⁻¹) could be calculated based on the binding constants. Moreover, the enthalpy and entropy changes are consistent with the “Enthalpy-Entropy Compensation” equation obtained from our previous work. The negative enthalpy and positive entropy indicated that the electrostatic interactions played a major role in DNP-BSA binding process. Site marker competitive displacement experiments were carried out by using fluorescence and isothermal titration calorimetry (ITC) methods. These results showed that DNP bound with high affinity to Sudlow’s site I (subdomain IIA) of BSA. The distance (r = 3.78 nm) between donor (BSA) and acceptor (DNP) was obtained according to the mechanism of fluorescence resonance energy transfer (FRET). Furthermore, the results of synchronous fluorescence and circular dichroism (CD) spectroscopic studies indicated that the microenvironment and the secondary conformation of BSA were altered. The above results were supported by theoretical molecular modeling methods.     

Study on the Fluorescence Quenching Reaction of Amitriptyline and Clomipramine Hydrochlorides with Eosin Y and its Analytical Application

Amitriptyline.HCl (AMI) and clomipramine.HCl (CMI) react with eosin Y (EY) in pH 3.8 NaAc-AcH buffer solution to form ion association complex which results in quenching of fluorescence of EY and appearance of a new resonance Rayleigh scattering (RSS) spectrum at 620 nm. The spectral characteristics of absorption, fluorescence and RSS spectra have been investigated. The factors influencing the reaction were studied and optimum conditions for the reaction have been determined. Based on fluorescence quenching, a simple and sensitive spectrofluorimetric method for determination of AMI and CMI has been developed. The fluorescence quenching intensity was measured at 550 nm using an excitation wavelength of 310 nm. The calibration graph was found to be rectilinear in the range 0.08–2.0 μg?mL^?1 with detection limit of 0.017 μg?mL^?1 for AMI and 0.06–2.0 μg?mL^?1 with detection limit of 0.015 μg?mL^?1 for CMI. The method can be satisfactorily applied to the determination of AMI and CMI in tablets without interference from commonly occurring exicipients. The recovery and RSD values obtained indicate good accuracy and precision of the method. The mechanism of the reaction and fluorescence quenching has also been discussed.