Toward an Alternative Intrinsic Probe for Spectroscopic Characterization of a Protein

The intrinsic fluorescent amino acid tryptophan is the unanimous choice for the spectroscopic investigation of proteins. However, several complicacies in the interpretation of tryptophan fluorescence in a protein are inevitable and an alternative intrinsic protein probe is a longstanding demand. In this contribution, we report an electron-transfer reaction in a human transporter protein (HSA) cavity which causes the tryptophan residue (Trp214) to undergo chemical modification to form one of its metabolites kynurenine (Kyn214). Structural integrity upon modification of the native protein is confirmed by dynamic light scattering (DLS) as well as near and far circular dichroism (CD) spectroscopy. Femtosecond-resolved fluorescence transients of the modified protein describe the dynamics of solvent molecules in the protein cavity in both the native and denatured states. In order to establish general use of the probe, we have studied the dipolar interaction of Kyn214 with a surface-bound ligand (crystal violet, CV) of the protein. By using the sensitivity of FRET, we have determined the distance between Kyn214 (donor) and CV (acceptor). Our study is an attempt to explore an alternative intrinsic fluorescence probe for the spectroscopic investigation of a protein. In order to establish the efficacy of the modification technique we have converted the tryptophan residues of other proteins (bovine serum albumin, chymotrypsin and subtilisin Carlsberg) to kynurenine and confirmed their structural integrity. We have also shown that catalytic activity of the enzymes remains intact upon the modification

Interaction of an Antituberculosis Drug with a Nanoscopic Macromolecular Assembly: Temperature-Dependent Förster Resonance Energy Transfer Studies on Rifampicin in an Anionic Sodium Dodecyl Sulfate Micelle

In this contribution, we report studies on the nature of binding of a potent antituberculosis drug, Rifampicin (RF) with a model drug delivery system, sodium dodecyl sulfate (SDS) micelle. Temperature dependent dynamic light scattering (DLS), conductometry, and circular dichroism (CD) spectroscopy have been employed to study the binding interaction of the drug with the micelle. The absorption spectrum of the drug RF in the visible region has been employed to study Förster resonance energy transfer (FRET) from another fluorescent drug Hoechst 33258 (H33258), bound to the micelle. Picosecond-resolved FRET studies at room temperature confirm the simultaneous binding of the two drugs to the micelle and the distance between the donor−acceptor pair is found to be 34 Å. The temperature dependent FRET study also confirms that the location and efficiency of drug binding to the micelle changes significantly at the elevated temperature. The energy transfer efficiency of the donor H33258, as measured from time-resolved studies, decreases significantly from 76% at 20 °C to 60% at 55 °C. This reveals detachment of some amount of the drug molecules from the micelles and increased donor−acceptor distance at elevated temperatures. The estimated donor−acceptor distance increases from a value of 33 Å at 20 °C to 37 Å at 55 °C. The picosecond resolved FRET studies on a synthesized DNA bound H33258 in RF solution have been performed to explore the interaction between the two. Our studies are expected to find relevance in the exploration of a potential vehicle for the vital drug rifampicin

Photoselective excited state dynamics in ZnO–Au nanocomposites and their implications in photocatalysis and dye-sensitized solar cells

Improving the performance of photoactive solid-state devices begins with systematic studies of the metal–semiconductor nanocomposites (NCs) upon which such devices are based. Here, we report the photo-dependent excitonic mechanism and the charge migration kinetics in a colloidal ZnO–Au NC system. By using a picosecond-resolved Förster resonance energy transfer (FRET) technique, we have demonstrated that excited ZnO nanoparticles (NPs) resonantly transfer visible optical radiation to the Au NPs, and the quenching of defect-mediated visible emission depends solely on the excitation level of the semiconductor. The role of the gold layer in promoting photolytic charge transfer, the activity of which is dependent upon the degree of excitation, was probed using methylene blue (MB) reduction at the semiconductor interface. Incident photon-to-current efficiency measurements show improved charge injection from a sensitizing dye to a semiconductor electrode in the presence of gold in the visible region. Furthermore, the short-circuit current density and the energy conversion efficiency of the ZnO–Au NP based dye-sensitized solar cell (DSSC) are much higher than those of a DSSC comprised of only ZnO NP. Our results represent a new paradigm for understanding the mechanism of defect-state passivation and photolytic activity of the metal component in metal–semiconductor nanocomposite systems

Ultrafast dynamics of excitons in semiconductor quantum dots on a plasmonically active nano-structured silver film

The excited state dynamics of core-shell type semiconductor quantum dots (QDs) of various sizes in close contact with a plasmonically active silver thin film has been demonstrated by using picosecond resolved fluorescence spectroscopy. The non-radiative energy transfer from the QDs to the metal surface is found to be of Föster resonance energy transfer (FRET) type rather than the widely expected nano-surface energy transfer (NSET) type. The slower rate of energy transfer processes compared to that of the electron transfer from the excited QDs to an organic molecule benzoquinone reveals an insignificant possibility of charge migration from the QDs to the metallic film

Functionalization of manganite nanoparticles and their interaction with biologically relevant small ligands: picosecond time-resolved FRET studies

We report molecular functionalization of the promising manganite nanoparticles La0.67Sr0.33MnO3 (LSMO) for their solubilization in aqueous environments. The functionalization of individual NPs with the biocompatible citrate ligand, as confirmed by Fourier transform infrared (FTIR) spectroscopy, reveals that citrates are covalently attached to the surface of the NPs. UV-VIS spectroscopic studies on the citrate functionalized NPs reveals an optical band in the visible region. Uniform size selectivity (2.6 nm) of the functionalization process is confirmed from high resolution transmission electron microscope (HRTEM). In the present study we have used the optical band of the functionalized NPs to monitor their interaction with other biologically important ligands. Forster resonance energy transfer (FRET) of a covalently attached probe 4-nitrophenylanthranilate (NPA) with the capped NPs confirm the attachment of the NPA ligands to the surface functional group (-OH) of the citrate ligand. The FRET of a DNA base mimic, 2-aminopurine (2AP), with the NPs confirms the surface adsorption of 2AP. Our study may find relevance in the study of the interaction of individual manganite NPs with drug/ligand molecules

Dynamics of light harvesting in ZnO nanoparticles

We have explored light harvesting of the complex of ZnO nanoparticles with the biological probe Oxazine 1 in the near-infrared region using picosecond-time-resolved fluorescence decay studies. We have used ZnO nanoparticles and Oxazine 1 as a model donor and acceptor, respectively, to explore the efficacy of the Förster resonance energy transfer (FRET) in the nanoparticle-dye system. It has been shown that FRET from the states localized near the surface and those in the bulk of the ZnO nanoparticles can be resolved by measuring the resonance efficiency for various wavelengths of the emission spectrum. It has been observed that the states located near the surface for the nanoparticles (contributing to visible emission at λ≈550 nm) can contribute to very high efficiency (>90%) FRET. The efficiency of light harvesting dynamics of the ZnO nanorods has also been explored in this study and they were found to have much less efficiency (∼40%) for energy transfer compared to the nanoparticles. The possibility of an electron transfer reaction has been ruled out from the picosecond-resolved fluorescence decay of the acceptor dye at the ZnO surface

Simultaneous binding of anti-tuberculosis and anti-thrombosis drugs to a human transporter protein: A FRET study

Although rifampicin (Rf) is one of the most effective antibiotics against infection caused by Mycobacterium tuberculosis, interaction of the drug with universal carrier protein in human blood plasma is not fully understood. Reduction of medicinal efficacy of other drugs, including anti-thrombosis drug warfarin (Wf), to the patients on Rf therapy also needs molecular understanding. In the present work we have studied interaction of Rf with one of the model carrier protein (human serum albumin). By using circular dichroism (CD) spectroscopy we have characterized the change in the secondary structure of the protein. The consequence of the simultaneous binding of the two drugs, Rf and Wf, on the structure of the protein has also been explored. Picosecond resolved Förster resonance energy transfer (FRET) from Wf to Rf explores possible binding sites of the anti-tuberculosis drug on the protein. In this report, we have discussed the potential problem of using the single tryptophan of the protein (Trp 214) as energy donor in FRET experiment for the characterization of the binding site of the drug Rf on the protein

Protein-Mediated Synthesis of Nanosized Mn-Doped ZnS: A Multifunctional, UV-Durable Bio-Nanocomposite

The design of synthetic nanoparticles (NPs) capable of recognizing given chemical entities in a specific and predictable manner is of great fundamental and practical importance. Herein, we report a simple, fast, water-soluble, and green phosphine free colloidal synthesis route for the preparation of multifunctional enzyme-capped ZnS bionanocomposites (BNCs) with/without transitional metal-ion doping. The enzymes α-Chymotrypsin (CHT), associated with the NPs, are demonstrated as an effectual host for organic dye Methylene Blue (MB) revealing the molecular recognition of such dye molecules by the BNCs. An effective hosting of MB in the close proximity of ZnS NPs (with ∼3 nm size) leads to photocatalysis of the dyes which has further been investigated with doped-semiconductors. The NP-associated enzyme α-CHT is found to be active toward a substrate (Ala-Ala-Phe-7-amido-4-methyl-coumarin), hence leads to significant enzyme catalysis. Irradiation induced luminescence enhancement (IILE) measurements on the BNCs clearly interpret the role of surface capping agents which protect against deep UV damaging of ZnS NPs

Ultrafast dynamics of excitons in semiconductor quantum dots on a plasmonically active nano-structured silver film

The excited state dynamics of core–shell type semiconductor quantum dots (QDs) of various sizes in close contact with a plasmonically active silver thin film has been demonstrated by using picosecond resolved fluorescence spectroscopy. The non-radiative energy transfer from the QDs to the metal surface is found to be of Förster resonance energy transfer (FRET) type rather than the widely expected nano-surface energy transfer (NSET) type. The slower rate of energy transfer processes compared to that of the electron transfer from the excited QDs to an organic molecule benzoquinone reveals an insignificant possibility of charge migration from the QDs to the metallic film