Internal Mechanical Response of a Polymer in Solution

Abstract

We observed single molecules of fluorescently labeled double-stranded (ds) λ DNA held in an antiBrownian electrokinetic trap. From the measured density fluctuations we extract the density-density response function of the molecule over times >4.5 ms and distances >250 nm, i.e., how a perturbation in density in one part of the molecule propagates through the rest of the molecule. We find a nonmonotonic radial dependence of the relaxation time. In contrast with earlier measurements on freely diffusing dsDNA, we observe clear signs of internal hydrodynamic interactions.Thermal fluctuations agitate molecules in solution over a broad range of times and distances. By passively watching the shape fluctuations of a thermally driven biopolymer, one can infer properties of the underlying interactions that determine the motion.Dynamic light scattering (DLS) and fluorescence correlation spectroscopy (FCS) each probe specific low-dimensional correlation functions of the shape fluctuations; these can be compared to predictions of Rouse, Zimm, or other hydrodynamic models [1]. The two techniques are complementary in that DLS probes fluctuations at a fixed k vector and FCS at a fixed position vector. Both techniques are also limited in the time scales they can probe: very fast fluctuations occur over distances too short to be detected optically, and very slow fluctuations are masked by translational diffusion.DLS has been used to measure the few longest relaxation times of DNA, and much theory has been devoted to predicting scattering functions for different polymer models [2,3]. FCS has also been used to probe the dynamic fluctuations of fluorescently labeled DNA [4,5]. Recently Shusterman and co-workers [6] probed the dynamics of the end monomer in single-stranded (ss) and double-stranded (ds) DNA using FCS. They observed Zimm dynamics in the ssDNA, but, surprisingly, Rouse dynamics in the dsDNA on time scales <10 ms. In contrast, the translational diffusion of dsDNA obeys Zimm scaling to high accuracy [7]. Shusterman et al. argued that a semiflexible partially draining polymer should show Rouse dynamics on short time scales and Zimm dynamics on long time scales. However, the observation times required to detect Zimm dynamics in dsDNA are longer than can be achieved with FCS.Here we directly measure the underlying time-dependent conformations of individual dsDNA molecules over times >4.5 ms and distances >250 nm. FCS-like and DLS-like correlation functions are special cases of the density-density covariance obtained from our data. Our experiment lacks the short-time resolution of DLS and FCS, but probes long-time dynamics (milliseconds to seconds) that are masked by translational diffusion in the other techniques. The data presented here provide the first detailed spatial picture of the fluctuating dynamics within a single conformationally relaxed polymer molecule. To study a single molecule in equilibrium, one would like to eliminate the motion of the center of mass (c.m.), without affecting internal motions. We used ...

Density fluctuations dispersion relationship for a polymer confined to a nanotube

“…5,6 In particular, for principal components ͑PC͒ of fluctuation of free DNA good agreement of experiment and theory was obtained while it was also established that the dynamics of these PC are not those of Zimm modes. [7][8][9] The work in nanochannels builds on a growing set of dynamic experimental studies of DNA in nanochannels, 10,11 and recent insights gained through simulation and theory. 12,13 We will report the observation of Rouse modes in the nanochannel system.…”

Structure–Fluorescence Contrast Relationship in Cyanine DNA Intercalators: Toward Rational Dye Design

“…[6] These attractive characteristics enabled DNA to be detected for the first time at a sensitivity comparable to that of radioactive probes, but without the danger inherent in radioactivity. [7] Although some sequence specificity that might lead to inhomogeneous staining has been demonstrated for both TOTO and, to a lesser extent, YOYO, [8][9][10] these cyanines and some derivatives have been used as general DNA stains in numerous DNA detection and quantitation assays, [11] such as the polymerase chain reaction, [12,13] DNA staining and fragment sizing, [14][15][16][17][18] DNA damage detection, [19,20] flow cytometry, [16,21,22] evaluation of biological activity, [23,24] DNA imaging [25][26][27][28] and DNA photocleavage. [29][30][31] YO and TO have also been covalently linked to oligonucleotides and inserted into peptide nucleic acids constructs, which become fluorescent upon hybridisation of the light-up probe to a specific complementary strand.…”

Real-time calibration of a feedback trap

“…First, it has been used to probe the physical or chemical properties of the trapped object. These include the diffusion constant and electric-field mobility of single particles, 5 their fluctuations, 6 and elastic and dissipation parameters that characterize the internal degrees of freedom of more complex objects. 7,8 Feedback traps can also be used to estimate the chemical properties of single molecules, including photodynamic and enzymatic properties of biomolecules 9 and the interplay between fluorescence spectroscopy and conformation at the single-molecule level.…”