Multi-spectroscopic investigation of the binding interaction of fosfomycin with bovine serum albumin.
ABSTRACT: The interaction between fosfomycin (FOS) and bovine serum albumin (BSA) has been investigated effectively by multi-spectroscopic techniques under physiological pH 7.4. FOS quenched the intrinsic fluorescence of BSA via static quenching. The number of binding sites n and observed binding constant KA were measured by the fluorescence quenching method. The thermodynamic parameters ΔG0, ΔH0 and ΔS0 were calculated at different temperatures according to the van't Hoff equation. The site of binding of FOS in the protein was proposed to be Sudlow's site I based on displacement experiments using site markers viz. warfarin, ibuprofen and digitoxin. The distance r between the donor (BSA) and acceptor (FOS) molecules was obtained according to the Förster theory. The effect of FOS on the conformation of BSA was analyzed using synchronous fluorescence spectra (SFS), circular dichroism (CD) and 3D fluorescence spectra. A molecular modeling study further confirmed the binding mode obtained by the experimental studies.
Project description:Introduction: Ascorbyl palmitate (AP) is an example of natural secondary food antioxidant, which has been used for oxidative rancidity prevention in food industry. In this study, the interaction of AP with bovine serum albumin (BSA) was investigated. Methods: The mechanism of BSA interaction with AP was investigated using spectroscopic methods (UV-Vis, fluorescence). The thermodynamic parameters including enthalpy change (ΔH), entropy change (ΔS), and Gibb's free energy (ΔG) were calculated using Van't Hoff equation at different temperatures. Results: The experimental results showed that UV-Vis absorption spectra of BSA decreased upon increasing AP concentration, indicating that the AP can bind to BSA. Formation of the AP-BSA complex was approved by quenching of fluorescence and the quenching mechanism was found to be resultant from dynamic procedure. The positive values of both ΔH and ΔS showed that hydrophobic forces were the major binding forces. The negative value of ΔG demonstrated that AP interacts with BSA spontaneously. Molecular docking results confirmed that AP binds to BSA through hydrophobic forces. Conclusion: The attained results showed that AP can bind to BSA and effectively distributed into the bloodstream.
Project description:In this report, the interaction between a phenanthrene-pyrene-based fluorescent probe (PPI) and bovine serum albumin (BSA), a transport protein, has been explored by steady-state emission spectroscopy, fluorescence anisotropy, far-ultraviolet circular dichroism (CD), time-resolved spectral measurements, and molecular docking simulation study. The blue shift along with emission enhancement indicates the interaction between PPI and BSA. The binding of the probe causes quenching of BSA fluorescence through both static and dynamic quenching mechanisms, revealing a 1:1 interaction, as delineated from Benesi-Hildebrand plot, with a binding constant of ?105 M-1, which is in excellent agreement with the binding constant extracted from fluorescence anisotropy measurements. The thermodynamic parameters, ?H°, ?S°, and ?G°, as determined from van't Hoff relationship indicate the predominance of van der Waals/extensive hydrogen-bonding interactions for the binding phenomenon. The molecular docking and site-selective binding studies reveal the predominant binding of PPI in subdomain IIA of BSA. From the fluorescence resonance energy transfer study, the average distance between tryptophan 213 of the BSA donor and the PPI acceptor is found to be 3.04 nm. CD study demonstrates the reduction of ?-helical content of BSA protein on binding with PPI, clearly indicating the change of conformation of BSA.
Project description:A synthesized and promising biologically hypoglycemic compound 5,6-Dichloro-2-[2-(pyridin-2-yl)ethyl]isoindoline-1,3-dione (5e) was studied for its binding to a model protein (bovine serum albumin; BSA) by spectroscopic and molecular simulation approaches. Fluorescence studies revealed that 5e quenched BSA's intrinsic fluorescence by static quenching. The experiments were performed at three different temperatures and the quenching constants and binding constants were evaluated. Stern-Volmer constant (Ksv) values decreased from 1.36?×?104 to 1.20?×?104 as the temperature increased suggesting static quenching involvement in the interaction. Decreased binding constants from 1.70?×?104 to 4.57?×?103 at higher temperatures indicated instability of the complex at rising temperatures. Site I (subdomain IIA) of BSA was found to interact with 5e. The thermodynamic results showed the binding interaction was spontaneous and enthalpy driven. The secondary structure alterations in BSA due to interaction with 5e were studied by UV-visible, synchronous fluorescence, and three-dimensional fluorescence spectra. The results indicate the 5e binds effectively to the BSA and thus, this study can be useful in further exploring the pharmacokinetics and pharmacodynamics of 5e.
Project description:Interaction of procainamide hydrochloride (PAH) with human serum albumin (HSA) is of great significance in understanding the pharmacokinetic and pharmacodynamic mechanisms of the drug. Multi-spectroscopic techniques were used to investigate the binding mode of PAH to HSA and results revealed the presence of static type of quenching mechanism. The number of binding sites, binding constants and thermodynamic parameters were calculated. The results showed a spontaneous binding of PAH to HSA and hydrophobic interactions played a major role. In addition, the distance between PAH and the Trp-214 was estimated employing the Förster's theory. Site marker competitive experiments indicated that the binding of PAH to HSA primarily took place in subdomain IIA (Sudlow's site I). The influence of interference of some common metal ions on the binding of PAH to HSA was studied. Synchronous fluorescence spectra (SFS), 3D fluorescence spectra and circular dichroism (CD) results indicated the conformational changes in the structure of HSA.
Project description:Repaglinide (RPG) regulates the amount of glucose by stimulating the pancreas to release insulin in the blood. In view of its biological importance, we have examined the interaction between RPG and a model protein, bovine serum albumin (BSA) employing various spectroscopic, electrochemical and molecular docking methods. Fluorescence spectra of BSA were recorded in the presence and absence of RPG in phosphate buffer of pH 7.4. Fluorescence intensity of BSA was decreased upon the addition of increased concentrations of RPG, indicating the interaction between RPG and BSA. Stern-Volmer quenching analysis results revealed that RPG quenched the intensity of BSA through dynamic quenching mechanism. This was further confirmed from the time-resolved fluorescence measurements. The binding constant as calculated from the spectroscopic and voltammetric results was observed to be in the order of 104?M-1?at 298?K, suggesting the moderate binding affinity between RPG and BSA. Competitive experimental results revealed that the primary binding site for RPG on BSA was site II. Absorption and circular dichroism studies indicated the changes in the secondary structure of BSA upon its interaction with RPG. Molecular simulation studies pointed out that RPG was bound to BSA in the hydrophobic pocket of site II.
Project description:Sodium benzoate (SB) is widely used as a preservative in food industry, and bovine serum albumin (BSA) is a major carrier protein similar to human serum albumin (HSA), the study of the binding between the two has great significance on human health. In this paper, we systematically investigated the binding of SB and BSA under the simulated physiological conditions combining with various common analytical methods, e.g., fluorescence, UV-vis absorption, synchronous fluorescence and circular dichroism (CD) spectra, as well as molecular docking method. The fluorescence quenching measurements were respectively carried out at 298 K, 303 K and 308 K using the Stern-Volmer method. The results reveal that ground state SB-BSA complex was formed within the binding constants from 2.02?×?104 to 7.9?×?103 M-1. Meanwhile, the negative values of ?H 0 (-?43.92 kJ mol-1) and ?S 0 (-?111.6 J mol-1 K-1) demonstrated that both the hydrogen binding interaction and van der Waals forces contributed to stabilizing the SB-BSA complex. The site marker competitive experiments show that the SB and BSA bound at site I. Furthermore, the experimental results of UV-vis absorption, synchronous fluorescence and CD spectra indicate that the binding of SB and BSA may change the conformation of BSA. In addition, the molecular docking experiment suggests that hydrogen bond was formed in the interaction between SB and BSA.
Project description:The interaction of afatinib (AFB) with bovine serum albumin (BSA) was examined via fluorescence and UV-Vis spectroscopy. Spectrofluorimetric measurements revealed that AFB can strongly quench the BSA intrinsic fluorescence through producing a non-fluorescent complex. This quenching mechanism was thoroughly investigated with regard to the type of quenching, binding constant, number of binding locations and the fundamental thermodynamic parameters. Subsequently, the association constant of AFB with BSA was computed at three different temperatures and was found to range from 7.34 to 13.19 x10(5) L mol(-1). Thermodynamic parameters calculations demonstrated a positive ?S? value with both negative ?H? and ?G? values for AFB-BSA complex, which in turn infers that a spontaneous binding is taking place with both electrostatic bonding and hydrophobic interactions participating in the binding of AFB and BSA. Similarly, the UV absorption spectra of AFB-BSA system were studied and confirmed the interaction. Conformational alteration of the protein upon binding to AFB was elaborated with the aid of three dimensional fluorescence measurements as well as synchronous fluorescence spectra.
Project description:Triptolide is a major active constituent isolated from Tripterygiumwilfordii Hook F, a Chinese herbal medicine. This study investigated the intermolecular interaction between triptolide and bovine serum albumin (BSA).The fluorescence, circular dichroism (CD) and molecular docking methods were used to investigate the intermolecular interaction between triptolide and BSA. The binding constant, the number of binding sites, binding subdomain and the thermodynamic parameters were measured.The results of this experiment revealed that the intrinsic fluorescence of BSA was effectively quenched by triptolide via static quenching. The experimental results of synchronous fluorescence and CD spectra showed that the conformation of BSA was changed in the presence of triptolide.It indicated that triptolide could spontaneously bind on site II (subdomain IIIA) of BSA mainly via hydrogen bonding interactions and Van der Waals force.
Project description:Recognition of elements of protein tertiary structure is crucial for biotechnological and biomedical tasks; this makes the development of optical sensors for certain protein surface elements important. Herein, we demonstrated the ability of iron(II) clathrochelates (<b>1</b>-<b>3</b>) functionalized with mono-, di- and hexa-carboxyalkylsulfide to induce selective circular dichroism (CD) response upon binding to globular proteins. Thus, inherently CD-silent clathrochelates revealed selective inducing of CD spectra when binding to human serum albumin (HSA) (<b>1</b>, <b>2</b>), beta-lactoglobuline (<b>2</b>) and bovine serum albumin (BSA) (<b>3</b>). Hence, functionalization of iron(II) clathrochelates with the carboxyalkylsulfide group appears to be a promising tool for the design of CD-probes sensitive to certain surface elements of proteins tertiary structure. Additionally, interaction of <b>1</b>-<b>3</b> with proteins was also studied by isothermal titration calorimetry, protein fluorescence quenching, electrospray ionization mass spectrometry (ESI-MS) and computer simulations. Formation of both 1:1 and 1:2 assemblies of HSA with <b>1</b>-<b>3</b> was evidenced by ESI-MS. A protein fluorescence quenching study suggests that <b>3</b> binds with both BSA and HSA via the sites close to Trp residues. Molecular docking calculations indicate that for both BSA and HSA, binding of <b>3</b> to Site I and to an "additional site" is more favorable energetically than binding to Site II.
Project description:Chrysin (5,7-dihydroxyflavone) is a flavonoid aglycone, which is found in nature and in several dietary supplements. During the biotransformation of chrysin, its conjugated metabolites chrysin-7-sulfate (C7S) and chrysin-7-glucuronide (C7G) are formed. Despite the fact that these conjugates appear in the circulation at much higher concentrations than chrysin, their interactions with serum albumin have not been reported. In this study, the complex formation of chrysin, C7S, and C7G with human (HSA) and bovine (BSA) serum albumins was investigated employing fluorescence spectroscopic, ultrafiltration, and modeling studies. Our major observations/conclusions are as follows: (1) Compared to chrysin, C7S binds with a threefold higher affinity to HSA, while C7G binds with a threefold lower affinity; (2) the albumin-binding of chrysin, C7S, and C7G did not show any large species differences regarding HSA and BSA; (3) tested flavonoids likely occupy Sudlow's Site I in HSA; (4) C7S causes significant displacement of Sudlow's Site I ligands, exerting an even stronger displacing ability than the parent compound chrysin. Considering the above-listed observations, the high intake of chrysin (e.g., through the consumption of dietary supplements with high chrysin contents) may interfere with the albumin-binding of several drugs, mainly due to the strong interaction of C7S with HSA.