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Binding interaction of a novel fluorophore with serum albumins: steady state fluorescence perturbation and molecular modeling analysis.

Pal U, Pramanik SK, Bhattacharya B, Banerji B, Maiti NC - Springerplus (2015)

Bottom Line: The fluorescence yield of the compound substantially increased inside hydrophobic protein surface and ~30 nm decrease in Stokes' shift, compared to aqueous solution, was observed.Thus, the molecule appears as a new fluorescence probe to report the nature of its binding site in terms of increased fluorescence quantum yield and decreased Stokes' shift.Further it could be useful to detect and study the drug binding site of specific protein of interest.

View Article: PubMed Central - PubMed

Affiliation: Structural Biology and Bioinformatics Division, Council of Scientific and Industrial Research (CSIR)-Indian Institute of Chemical Biology (IICB), Kolkata, West Bengal India.

ABSTRACT
Fluorescence emission and anisotropy are widely used to measure the binding parameters and kinetic behavior of reactions that cause a change in the rotational time of a fluorescent molecule. We report here fluorescence emission and anisotropy behavior of a newly synthesized novel naphthalene base fluorophore (methyl 3-[(6-{[2-(tert-butoxy)-2-oxoethyl] (4-methoxyphenyl)amino}naphthalen-2-yl)formamido]propanoate) in several solution conditions including its binding to human and bovine serum albumin proteins both in their native and denatured states. The fluorescence yield of the compound substantially increased inside hydrophobic protein surface and ~30 nm decrease in Stokes' shift, compared to aqueous solution, was observed. Shift in fluorescence excitation peak position from the absorption peak of the molecule was ~8 nm in protein solution. This indicated possible alteration of excited state geometry of the compound by the globular fold of albumins. In addition, we measured the steady state fluorescence anisotropy of the molecule to evaluate several thermodynamic parameters and the results suggested the binding was energetically favorable. The measured ΔG° was ~-30 kJ mol(-1) and the derived dissociation constant was ~10(-6) M. The molecular docking analysis further highlighted the nonspecific association of the compound with the proteins and hydrophobic forces may have a significant role in the binding processes. Under the denatured condition of the protein, the compound lost its binding efficacy and reduction in fluorescence intensity was observed. Thus, the molecule appears as a new fluorescence probe to report the nature of its binding site in terms of increased fluorescence quantum yield and decreased Stokes' shift. It can also report the changes in the binding site due to global change in protein structure such as unfolding/misfolding often linked to several human disorder. Further it could be useful to detect and study the drug binding site of specific protein of interest.

No MeSH data available.


Related in: MedlinePlus

Fluorescence emission and anisotropy change of compound 5 in presence of serum albumin. Compound 5 concentration was kept constant at 0.5 μM and the protein concentration was varied from 0 through 5.5 μM. a The changes in fluorescence emission spectrum of compound 5 as a function of BSA concentration. b The changes in compound 5 fluorescence anisotropy with increasing concentration of BSA and the fitted Langmuir isotherm
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Fig2: Fluorescence emission and anisotropy change of compound 5 in presence of serum albumin. Compound 5 concentration was kept constant at 0.5 μM and the protein concentration was varied from 0 through 5.5 μM. a The changes in fluorescence emission spectrum of compound 5 as a function of BSA concentration. b The changes in compound 5 fluorescence anisotropy with increasing concentration of BSA and the fitted Langmuir isotherm

Mentions: Compound 5 absorbs UV–visible light strongly at the wavelengths where protein absorbs. It also has a strong absorption band in the fluorescence emission range of protein. Therefore, protein intrinsic fluorescence perturbation with compound 5 or the energy transfer from protein to compound 5 is not an eminent choice to probe the binding interactions of compound 5 with proteins. However, the changes in compound 5 fluorescence may be monitored as a parameter of binding. With the increasing concentration of serum albumins, we have found that the fluorescence intensity of the compound 5 increases (Fig. 2a, Additional file 1: Figure S1A). A blue shift in the emission maximum was also observed. It indicated compound 5 binding to the hydrophobic grooves of serum albumins. Such binding causes solvent exclusion of compound 5 and the energy that is otherwise spent in solvent relaxation, is gained by the emitting photons. Here, about 30 nm decrease in the Stokes’ shift in compound 5 fluorescence corresponds to ~0.2 eV energy gain by each emitted photon.


Binding interaction of a novel fluorophore with serum albumins: steady state fluorescence perturbation and molecular modeling analysis.

Pal U, Pramanik SK, Bhattacharya B, Banerji B, Maiti NC - Springerplus (2015)

Fluorescence emission and anisotropy change of compound 5 in presence of serum albumin. Compound 5 concentration was kept constant at 0.5 μM and the protein concentration was varied from 0 through 5.5 μM. a The changes in fluorescence emission spectrum of compound 5 as a function of BSA concentration. b The changes in compound 5 fluorescence anisotropy with increasing concentration of BSA and the fitted Langmuir isotherm
© Copyright Policy - OpenAccess
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4582037&req=5

Fig2: Fluorescence emission and anisotropy change of compound 5 in presence of serum albumin. Compound 5 concentration was kept constant at 0.5 μM and the protein concentration was varied from 0 through 5.5 μM. a The changes in fluorescence emission spectrum of compound 5 as a function of BSA concentration. b The changes in compound 5 fluorescence anisotropy with increasing concentration of BSA and the fitted Langmuir isotherm
Mentions: Compound 5 absorbs UV–visible light strongly at the wavelengths where protein absorbs. It also has a strong absorption band in the fluorescence emission range of protein. Therefore, protein intrinsic fluorescence perturbation with compound 5 or the energy transfer from protein to compound 5 is not an eminent choice to probe the binding interactions of compound 5 with proteins. However, the changes in compound 5 fluorescence may be monitored as a parameter of binding. With the increasing concentration of serum albumins, we have found that the fluorescence intensity of the compound 5 increases (Fig. 2a, Additional file 1: Figure S1A). A blue shift in the emission maximum was also observed. It indicated compound 5 binding to the hydrophobic grooves of serum albumins. Such binding causes solvent exclusion of compound 5 and the energy that is otherwise spent in solvent relaxation, is gained by the emitting photons. Here, about 30 nm decrease in the Stokes’ shift in compound 5 fluorescence corresponds to ~0.2 eV energy gain by each emitted photon.

Bottom Line: The fluorescence yield of the compound substantially increased inside hydrophobic protein surface and ~30 nm decrease in Stokes' shift, compared to aqueous solution, was observed.Thus, the molecule appears as a new fluorescence probe to report the nature of its binding site in terms of increased fluorescence quantum yield and decreased Stokes' shift.Further it could be useful to detect and study the drug binding site of specific protein of interest.

View Article: PubMed Central - PubMed

Affiliation: Structural Biology and Bioinformatics Division, Council of Scientific and Industrial Research (CSIR)-Indian Institute of Chemical Biology (IICB), Kolkata, West Bengal India.

ABSTRACT
Fluorescence emission and anisotropy are widely used to measure the binding parameters and kinetic behavior of reactions that cause a change in the rotational time of a fluorescent molecule. We report here fluorescence emission and anisotropy behavior of a newly synthesized novel naphthalene base fluorophore (methyl 3-[(6-{[2-(tert-butoxy)-2-oxoethyl] (4-methoxyphenyl)amino}naphthalen-2-yl)formamido]propanoate) in several solution conditions including its binding to human and bovine serum albumin proteins both in their native and denatured states. The fluorescence yield of the compound substantially increased inside hydrophobic protein surface and ~30 nm decrease in Stokes' shift, compared to aqueous solution, was observed. Shift in fluorescence excitation peak position from the absorption peak of the molecule was ~8 nm in protein solution. This indicated possible alteration of excited state geometry of the compound by the globular fold of albumins. In addition, we measured the steady state fluorescence anisotropy of the molecule to evaluate several thermodynamic parameters and the results suggested the binding was energetically favorable. The measured ΔG° was ~-30 kJ mol(-1) and the derived dissociation constant was ~10(-6) M. The molecular docking analysis further highlighted the nonspecific association of the compound with the proteins and hydrophobic forces may have a significant role in the binding processes. Under the denatured condition of the protein, the compound lost its binding efficacy and reduction in fluorescence intensity was observed. Thus, the molecule appears as a new fluorescence probe to report the nature of its binding site in terms of increased fluorescence quantum yield and decreased Stokes' shift. It can also report the changes in the binding site due to global change in protein structure such as unfolding/misfolding often linked to several human disorder. Further it could be useful to detect and study the drug binding site of specific protein of interest.

No MeSH data available.


Related in: MedlinePlus