Time-resolved Fluorescence Stokes Shift Research Articles

Molecular Picture of the Effect of Cosolvent Crowding on Ligand Binding and Dispersed Solvation Dynamics in G-Quadruplex DNA.

Understanding molecular interactions and dynamics of proteins and DNA in a cell-like crowded environment is crucial for predicting their functions within the cell. Noncanonical G-quadruplex DNA (GqDNA) structures adopt various topologies that were shown to be strongly affected by molecular crowding. However, it is unknown how such crowding affects the solvation dynamics in GqDNA. Here, we study the effect of cosolvent (acetonitrile) crowding on ligand (DAPI) solvation dynamics within human telomeric antiparallel GqDNA through direct comparison of time-resolved fluorescence Stokes shift (TRFSS) experiments and molecular dynamics (MD) simulations results. We show that ligand binding affinity to GqDNA is drastically affected by acetonitrile (ACN). Solvation dynamics probed by DAPI in GqDNA groove show dispersed dynamics from ∼100 fs to 10 ns in the absence and presence of 20% and 40% (v/v) ACN. The nature of dynamics remain similar in buffer and 20% ACN, although in 40% ACN, distinct dynamics is observed in <100 ps. MD simulations performed on GqDNA/DAPI complex reveal preferential solvation of ligand by ACN, particularly in 40% ACN. Simulated solvation time-correlation functions calculated from MD trajectories compare very well to the overall solvation dynamics of DAPI in GqDNA, observed in experiments. Linear response decomposition of simulated solvation correlation functions unfolds the origin of dispersed dynamics, showing that the slower dynamics is dominated by DNA-motion in the presence of ACN (and also by the ACN dynamics at higher concentration). However, water-DNA coupled motion controls the slow dynamics in the absence of ACN. Our data, thus, unravel a detailed molecular picture showing that though ACN crowding affect ligand binding affinity to GqDNA significantly, the overall dispersed solvation dynamics in GqDNA remain similar in the absence and the presence of 20% ACN, albeit with a small effect on the dynamics in the presence of 40% ACN due to preferential solvation of ligand by ACN.

Origin of Slow Solvation Dynamics in DNA: DAPI in Minor Groove of Dickerson-Drew DNA.

The measurement and understanding of collective solvation dynamics in DNA have vital biological implications, as protein and ligand binding to DNA can be directly controlled by complex electrostatic interactions of anionic DNA and surrounding dipolar water, and ions. Time-resolved fluorescence Stokes shift (TRFSS) experiments revealed anomalously slow solvation dynamics in DNA much beyond 100 ps that follow either power-law or slow multiexponential decay over several nanoseconds. The origin of such dispersed dynamics remains difficult to understand. Here we compare results of TRFSS experiments to molecular dynamics (MD) simulations of well-known 4',6-diamidino-2-phenylindole (DAPI)/Dickerson-Drew DNA complex over five decades of time from 100 fs to 10 ns to understand the origin of such dispersed dynamics. We show that the solvation time-correlation function (TCF) calculated from 200 ns simulation trajectory (total 800 ns) captures most features of slow dynamics as measured in TRFSS experiments. Decomposition of TCF into individual components unravels that slow dynamics originating from dynamically coupled DNA-water motion, although contribution from coupled water-Na+ motion is non-negligible. The analysis of residence time of water molecules around the probe (DAPI) reveals broad distribution from ∼6 ps to ∼3.5 ns: Several (49 nos.) water molecules show residences time greater than 500 ps, of which at least 14 water molecules show residence times of more than 1 ns in the first solvation shell of DAPI. Most of these slow water molecules are found to occupy two hydration sites in the minor groove near DAPI binding site. The residence time of Na+, however, is found to vary within ∼17-120 ps. Remarkably, we find that freezing the DNA fluctuations in simulation eliminates slower dynamics beyond ∼100 ps, where water and Na+ dynamics become faster, although strong anticorrelation exists between them. These results indicate that primary origin of slow dynamics lies within the slow fluctuations of DNA parts that couple with nearby slow water and ions to control the dispersed collective solvation dynamics in DNA minor groove.

The Time-Resolved Fluorescence Stokes Shift of Cucurbit[6]Uril Complexes with a Pyridinium Styryl Dye.

The time-resolved Stokes shift of fluorescence of styryl dye, trans-4-[4-(dimethylamino)-styryl]-1-methylpyridinium iodide (DASPI), and its host-guest complexes with cucurbit[6]uril (CB[6]) was studied by up-conversion technique on a 5ps time scale. In all cases, the solvation time correlation function - that describes relaxation of the fluorescent state - contains two components. The time constant of the first component, which is related to the vibrational relaxation of the dye cation, increases from 118fs without CB[6] to 224fs in the presence CB[6]. It suggests the production of CB[6] complexes to disturb the hydrogen-bond net in the vicinity of DASPI cation. The time constant of the second component, which is associated with solvation, changes from 0.81ps without CB[6] to 1.15ps in the presence of the cavitand. The observed increase can be related to the displacement of the dye cation deeper into CB[6] that allows considering DASPI-CB[6] complexes as a light-controlled molecular machine with a response time on the picosecond scale.

Dynamics of water and ions around DNA: What is so special about them?

Water around biomolecules is special for behaving strangely - both in terms of structure and dynamics, while ions are found to control various interactions in biomolecules such as DNA, proteins and lipids. The questions that how water and ions around these biomolecules behave in terms of their structure and dynamics, and how they affect the biomolecular functions have triggered tremendous research activities worldwide. Such activities not only unfolded important static and dynamic properties of water and ions around these biomolecules, but also provoked heated debate regarding their explanation and role in biological functions. DNA, being negatively charged, interacts strongly with the surrounding dipolar water and positively charged counterions, leading to complex electrostatic coupling of water and ions with the DNA. Recent timeresolved fluorescence Stokes shift experiments and related computer simulation studies from our and other laboratories have unfolded some unique dynamic characteristics of water and ions near different structures of DNA. These results are discussed here to showcase the specialty of water and ion dynamics around DNA.

How Heterogeneous Are Trehalose/Glycerol Cryoprotectant Mixtures? A Combined Time-Resolved Fluorescence and Computer Simulation Investigation.

Heterogeneity and molecular motions in representative cryoprotectant mixtures made of trehalose and glycerol are investigated in the temperature range 298 ≤ T (K) ≤ 353, via time-resolved fluorescence Stokes shift and anisotropy measurements, and molecular dynamics simulations of four-point density-time correlations and H-bond relaxations. Mixtures containing 5 and 20 wt % of trehalose along with neat glycerol are studied. Viscosity coefficients for these systems lie in the range 0.30 < η (P) < 23. Measured solute (Coumarin 153) rotation and solvation times reveal a substantial departure from the hydrodynamic viscosity dependence, suggesting the strong microheterogeneous nature of these systems. Fluorescence anisotropy decays are highly nonexponential, reflecting a non-Markovian character of the medium friction. A complete missing of the Stokes shift dynamics in these systems at 298 K but partial detection of it at other higher temperatures (shift magnitude being ∼400-600 cm-1) indicates rigid solute environments. An amorphous solid-like feature emerges in the simulated radial distribution functions at these temperatures. Analyses of mean squared displacements reveal rattling-in-a-cage motion, non-Gaussian displacement distributions, and strong dynamic heterogeneity features. Simulated dynamic structure factors and four-point correlations hint, respectively, at very long α-relaxation and correlated time scales at 298 K. This explains the long solute rotation times (∼80-200 ns) measured at 298 K. Stretched exponential decay of the simulated H-bond relaxations with long time scales further highlights the strong temporal heterogeneity and slow dynamics inherent to these systems. In summary, this work provides the first insight into the molecular motions and interspecies interaction in a representative cryoprotectant mixture, and stimulates further study to investigate the interconnection between cryoprotection and dynamic heterogeneity.

Solvent sorting in (mixed solvent + electrolyte) systems: Time-resolved fluorescence measurements and theory

In this manuscript we explore electrolyte-induced modification of preferential solvation of a dipolar solute dissolved in a binary mixture of polar solvents. Composition dependence of solvation characteristics at a fixed electrolyte concentration has been followed. Binary mixtures of two different polarities have been employed to understand the competition between solute-ion and solute-solvent interactions. Time-resolved fluorescence Stokes shift and anisotropy have been measured for coumarin 153 (C153) in moderately polar (ethyl acetate + 1-propanol) and strongly polar (acetonitrile + propylene carbonate) binary mixtures at various mixture compositions, and in the corresponding 1.0 M solutions of LiClO4. Both the mixtures show red shifts in C153 absorption and fluorescence emission upon increase of mole fraction of the less polar solvent component in presence of the electrolyte. In addition, measured average solvation times become slower and rotation times faster for the above change in the mixture composition. A semi-molecular theory based on solution density fluctuations has been developed and found to successfully capture the essential features of the measured Stokes shift dynamics of these complex multi-component mixtures. Dynamic anisotropy results have been analyzed by using both Stokes-Einstein-Debye (SED) and Dote-Kivelson-Schwartz (DKS) theories. The importance of local solvent structure around the dissolved solute has been stressed.

Medium decoupling of dynamics at temperatures ∼100 K above glass-transition temperature: A case study with (acetamide + lithium bromide/nitrate) melts

Time-resolved fluorescence Stokes shift and anisotropy measurements using a solvation probe in [0.78CH(3)CONH(2) + 0.22{f LiBr + (1-f) LiNO(3)}] melts reveal a strong decoupling of medium dynamics from viscosity. Interestingly, this decoupling has been found to occur at temperatures ∼50-100 K above the glass transition temperatures of the above melt at various anion concentrations (f(LiBr)). The decoupling is reflected via the following fractional viscosity dependence (η) of the measured average solvation and rotation times (<τ(s)> and <τ(r)>, respectively): <τ(x)> ∝ (η∕T)(p) (x being solvation or rotation), with p covering the range, 0.20 < p < 0.70. Although this is very similar to what is known for deeply supercooled liquids, it is very surprising because of the temperature range at which the above decoupling occurs for these molten mixtures. The kinship to the supercooled liquids is further exhibited via p which is always larger for <τ(r)> than for <τ(s)>, indicating a sort of translation-rotation decoupling. Multiple probes have been used in steady state fluorescence measurements to explore the extent of static heterogeneity. Estimated experimental dynamic Stokes shift for coumarin 153 in these mixtures lies in the range, 1000 < Δν(t)/cm(-1) < 1700, and is in semi-quantitative agreement with predictions from our semi-molecular theory. The participation of the fluctuating density modes at various length-scales to the observed solvation times has also been investigated.

Probe Position Dependence of DNA Dynamics: Comparison of the Time-Resolved Stokes Shift of Groove-Bound to Base-Stacked Probes

Time-resolved fluorescence Stokes shift dynamics of a fluorescent probe, 4',6-diamidino-2-phenylindole (DAPI), inside the minor groove of the DNA is measured over five decades in time spans from 100 fs to 10 ns. Two different techniques, fluorescence up-conversion and time correlated single photon counting, are combined to obtain the time-resolved emission spectra of DAPI in DNA over the entire five decades in time. Having the dynamics of groove-bound DAPI in DNA measured over such a broad time window, we are able to convincingly compare our data to earlier time-resolved fluorescence results of a base-stacked probe that replaces a DNA base pair. Results show that the dynamics measured with either the groove-bound or the base-stacked probe are similar in the time span of 100 fs to approximately 100 ps but differ substantially from approximately 100 ps to 10 ns. Our present data also help to reconcile the previously reported molecular dynamics simulation results and provide important clues that the groove-bound water molecules inside DNA are mainly responsible for the slow dynamics seen in native DNA.

Characterization of the solvation dynamics of an ionic liquid via molecular dynamics simulation

The solvation dynamics of ionic liquids have been the subject of intense experimental study but remain poorly understood. We present the results of molecular dynamics simulations of the solvation dynamics of the ionic liquid 1-butyl-3-methylimidazolium hexafluorophosphate in response to photoexcitation of the fluorescent dye coumarin-153. We reproduce the time-resolved fluorescence Stokes shift using linear response theory, then use novel statistical techniques to analyze cation and anion contributions to the signal. We find that the solvation dynamics are dominated by collective ionic motion and characterize the time scale for various features of the collective response. Further, we use the Steele analysis [Mol. Phys. 61, 1031 (1987)] to characterize the contributions to the observed Stokes shift made by translational and rovibrational degrees of freedom. Our results indicate that in contrast to molecular liquids, the rovibrational response is trivial and the observed fluorescence response arises almost entirely from ionic translation. Our results resolve previously open questions in the literature about the nature of the rapid dynamics in room-temperature ionic liquids and offer insight into the physical principles governing ionic liquid behavior on longer time scales.

Comparison of time-resolved fluorescence Stokes shift measurements to a molecular theory of solvation dynamics

The results of time-resolved Stokes shift measurements are compared to a molecular theory of ionic solvation dynamics recently solved by Rips, Klafter, and Jortner [J. Chem. Phys. 88, 3246 (1988)]. Although the theoretical predictions show only semiquantitative agreement with experiment, this molecularly based model provides a satisfying framework for understanding the large departures observed from the behavior predicted by simple continuum treatments of solvation dynamics.

A time-resolved fluorescence study of the dynamic stokes shift of trans-4-dimethylamino-4'-cyanostilbene

The time-resolved fluorescence Stokes shift of trans-4-dimethylamino-4'-cyanostilbene has been measured as a function of temperature in polar solvents. Fluorescence spectra were found to exhibit exponential rise times of 80 to 640 ps between 297 and 250 K. These observations are attributed to two independent relaxation processes in alcohols: to reorientation of the solvent cage and to relaxation of the solute. The rate of the former process depends on the strength of specific solvent-solute interactions.

Picosecond dynamic stokes shift of α-naphthylamine

The time-resolved fluorescence Stokes shift of α-naphthylamine in polar solvents has been measured at room temperature using a mode-locked Nd: YAG laser and a streak camera. Fluorescence spectra show a red-shift with an exponential rise time of 52 ps in isopropanol at 295 K. The rate of the relaxation process involved in the Stokes shift appears to be dependent on the extent of the change in dipole moment of the solute upon excitation.