A spectral deciphering of the binding interaction of an intramolecular charge transfer fluorescence probe with a cationic protein: thermodynamic analysis of the binding phenomenon combined with blind docking study (original) (raw)
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Chemical Physics, 2008
The enhanced fluorescence and anisotropy of the ESIPT emission of 3-hydroxy-2-naphthoic acid (3HNA) in the complexes of 3HNA with bovine serum albumin (BSA) and human serum albumin (HSA) showed the formation of 1:1 complex [binding constant = 5.3 Â 10 5 M À1 for BSA and = 2.2 Â 10 5 M À1 for HSA]. The ESIPT emission of the probe in non-polar, polar protic, polar aprotic and mixed solvents indicate that the position and the quantum yield of the emission are governed by the intermolecular hydrogen bonding ability and the polarity/polarizability of the solvents. Rotational correlation time of 3HNA (2.4 ns and 5.2 ns in BSA and HSA, respectively) obtained from the time resolved anisotropy decay of the ESIPT emission suggests motional restriction of the probe. Docking studies reveal H-bonding of some residues with the probe and loss of accessible surface area of several residues located near binding site.
Journal of Photochemistry and Photobiology A: Chemistry, 2012
Binding interaction of plasma protein bovine serum albumin (BSA) with external flexible charge transfer fluorophore 5-(4-dimethylamino-phenyl)-penta-2,4-dienenitrile (DMAPPDN) has been explored at physiological pH (7.4) by steady state absorption, emission, fluorescence anisotropy, Red Edge Excitation Shift (REES), far-UV circular dichroism (CD), time resolved spectral measurements in combination with molecular docking and molecular dynamics (MD) simulation studies. Chemical denaturation of the protein bound probe by guanidine hydrochloride (GdnHCl) has also been tracked using the spectral response of DMAPPDN. Interaction of the probe with BSA is reflected by the massive blue shift of the fluorophore emission maxima (78 nm) with the enhancement of fluorescence intensity due to change of hydrophobic micro-environment of the probe compared to a little change in protein secondary structure. Benesi-Hildebrand plot reveals spontaneous formation of 1:1 BSA-DMAPPDN complex with binding constant 8.821 ± 0.01 × 10 3 M −1 and binding free energy change −5.359 kcal mol −1 . Molecular docking study supports the binding of probe in the hydrophobic cavity of sub domain IIA of BSA. The distance for energy transfer from tryptophan of BSA to DMAPPDN measured from fluorescence resonance energy transfer (FRET) results is in good agreement with results of molecular docking study. MD simulation predicts greater stability of BSA-DMAPPDN complex compared to the free protein.
Biopolymers, 2012
The nature of binding of specially designed charge transfer (CT) fluorophore at the hydrophobic protein interior of human serum albumin (HSA) has been explored by massive blue-shift (82 nm) of the polarity sensitive probe emission accompanying increase in emission intensity, fluorescence anisotropy, red edge excitation shift, and average fluorescence lifetimes. Thermal unfolding of the intramolecular CT probe bound HSA produces almost opposite spectral changes. The spectral responses of the molecule reveal that it can be used as an extrinsic fluorescent reporter for similar biological systems. Circular dichrosim spectra, molecular docking, and molecular dynamics simulation studies scrutinize this binding process and stability of the protein probe complex more closely. # 2012 Wiley Periodicals, Inc. Biopolymers 97: 766-777, 2012.
Photochemistry and Photobiology, 2010
The carcinogenic drug 4‐nitroquinoline‐1‐oxide (4NQO) has been found to bind with the protein hen egg white lysozyme as evident from fluorescence quenching experiments. The binding constant and stoichiometry have been determined. The values of the thermodynamic parameters indicate that the interaction is an enthalpy‐driven spontaneous phenomenon. The experimental value of change in free energy is similar to that obtained from the docking study. The far UV circular dichroism spectra show some changes in the secondary structure of protein. The high value of bimolecular quenching constant leads to the possibility of Förster resonance energy transfer (FRET). Along with FRET, the photoinduced electron transfer (PET) from tryptophan residue of protein to 4NQO has also been evident from the transient absorption spectra obtained in laser flash photolysis experiments. The simultaneous occurrence of FRET and PET is the key factor for quenching of intrinsic fluorescence of the protein as it bi...
Photochemical & Photobiological Sciences, 2012
Structural modification through binding interaction of plasma protein bovine serum albumin (BSA) with an extrinsic charge transfer fluorophore 5-(4-dimethylamino-phenyl)-penta-2,4-dienoic acid (DMAPPDA) and its response to external perturbation due to interactions with surfactant sodium dodecyl sulphate (SDS) have been explored at physiological pH by steady state absorption, emission, fluorescence anisotropy, red edge excitation shift, far-UV circular dichroism and time resolved spectral measurements in combination with Molecular Docking and Molecular Dynamics (MD) simulation. Interaction of the probe with BSA is reflected by a small change in protein secondary structure with fluorescence enhancement and blue shift of probe emission. Molecular docking studies revealed that the probe binds to the hydrophobic cavity of sub-domain IIA of BSA. The distance for energy transfer from the tryptophan of BSA to the bound DMAPPDA measured by Fluorescence Resonance Energy Transfer is in good agreement with the molecular docking results. MD simulation predicts stabilization of the complex with respect to the bare molecule. Interaction of BSA and SDS with DMAPPDA supports the movement of the probe from hydrophilic free water region to a more restricted hydrophobic zone inside the protein.
The Journal of Physical Chemistry, 1996
The photophysics of short linear peptides of general formula Boc-Leu-Leu-Lys(P)-(AA) n-Leu-Leu-Lys(N)-OtBu, where AA) Ala or Aib (R-aminoisobutyric acid), and P and N are protoporphyrin IX and naphthalene, respectively, covalently bound to-amino groups of lysine side-chains, were investigated in water/methanol 75/25 (v/v) solution by steady-state and time-resolved fluorescence experiments. Quenching of the excited naphthyl chromophore takes place by electronic energy transfer to the porphyrin ground state, and proceeds on a time scale of 3-8 ns. A minor and slower fluorescence lifetime measures the decay of the exciplexes. Quenching efficiencies exhibit a different trend, depending on whether AA) Ala or Aib, indicating differences in the structural features of the two series of peptides. Consistently, CD spectra suggest that the former compounds populate R-helical structures, while the latter ones possibly attain a 3 10-helix conformation, in agreement with the proven ability of Aib to form 3 10-helices in solution. The increased percentage of intramolecular H-bonds in the P(Aib) n N as compared to the corresponding P(Ala) n N peptides, as determined by IR spectra in dilute CD 3 OD or CDCl 3 solution, confirms this conclusion. The fluorescence results were satisfactorily described by a dipole-dipole interaction mechanism, provided the mutual orientations of N and P groups are taken into account, which implies that interconversion among conformational substates of chromophore linkage is slow on the time scale of the transfer process. Conformational statistics analysis shows a rather wide interprobe separation distance distribution for each peptide, owing to the aliphatic portion of the side-chains carrying the chromophores, but theoretical conformational analysis indicates that only a few energetically favored conformers are the major contributors to the energy transfer process.
Fluorescence spectroscopy as a tool to investigate protein interactions
Current Opinion in Biotechnology, 1997
Recent advances in the use of fluorescence spectroscopy to study protein interactions have primarily involved combinations of classic fluorescence techniques, novel probe and coupling chemistries, and advances in laser excitation and detection capabilities. For example, new coupling strategies for fluorescent probes have allowed the first determination of the AG" describing the insertion of a protein into a membrane. Fluorescently labeled oligonucleotides with specific protein-binding sequences have been used to study both protein-DNA associations and oligonucleotide hybridization using anisotropy changes. The first kinetic data describing a DNA-protein binding event was collected with stopped-flow fluorescence instrumentation. Combining scanning fluctuation correlation spectroscopy with a two-photon excitation source improved this technique so that it may now be used to study protein self-associations. Addresses Electronic identifier: 0958-l 669-008-00045 0 Current Biology Ltd ISSN 0958-l 669
Techniques in Protein Chemistry III, 1992
We used time-dependent fluorescence energy transfer to recover the site-to-site distance distributions and diffusion coefficients for a flexible donor-acceptor pair and for the proteins melittin (random coil and a-helix states) and troponin I (native and denatured). Energy transfer occurred from single tryptophan residues to dansyl acceptors. The differential equation describing the donor population in the presence of site-to-site diffusion (Biopolymers 17:11-31, 1978) was solved numerically.
The Journal of Physical Chemistry B, 2013
The structures, dynamics and energetics of the protonated, derivatized peptide DyeX-(Pro) 4 -Arg + -Trp, where "Dye" stands for the BODIPY analogue of tetramethylrhodamine and X is a (CH 2 ) 5 linker, have been investigated using a combination of modeling approaches in order to provide a numerical framework to the interpretation of fluorescence quenching data in the gas phase. Molecular dynamics (MD) calculations using the new generation AMOEBA force field were carried out using a representative set of conformations, at eight temperatures ranging from 150 to 500 K. Force field parameters were derived from ab initio calculations for the Dye. Strong electrostatic, polarization and dispersion interactions combine to shape this charged peptide. These effects arise in particular from the electric field generated by the charge of the protonated arginine and from several hydrogen bonds that can be established between the Dye linker and the terminal Trp. This conclusion is based on both the analysis of all structures generated in the MD simulations and on an energy decomposition analysis at classical and quantum mechanical levels. Structural analysis of the simulations at the different temperatures reveals that the relatively rigid polyproline segment allows for the Dye and Trp indole side chain to adopt stacking conformations favorable to electron transfer, yielding support to a model in which it is electron transfer from tryptophan to the dye that drives fluorescence quenching.