Polyelectrolyte properties of single stranded DNA measured using SAXS and single‐molecule FRET: Beyond the wormlike chain model

Abstract

Nucleic acids are highly charged polyelectrolytes that interact strongly with salt ions. Rigid, base-paired regions are successfully described with worm like chain models, but non base-paired single stranded regions have fundamentally different polymer properties because of their greater flexibility. Recently, attention has turned to single stranded nucleic acids due to the growing recognition of their biological importance, as well as the availability of sophisticated experimental techniques sensitive to the conformation of individual molecules. We investigate polyelectrolyte properties of poly(dT), an important and widely studied model system for flexible single stranded nucleic acids, in physiologically important mixed mono- and di-valent salt. We report measurements of the form factor and interparticle interactions using SAXS, end to end distances using smFRET, and number of excess ions using ASAXS. We present a coarse-grained model that accounts for flexibility, excluded volume, and electrostatic interactions in these systems. Predictions of the model are validated against experiment. We also discuss the state of all-atom, explicit solvent Molecular Dynamics simulations of poly(dT), the next step in understanding the complexities of ion interactions with these highly charged and flexible polymers.

Multi-enzyme complexes on DNA scaffolds capable of substrate channelling with an artificial swinging arm

“…Furthermore, titrating in free, unlabeled ssDNA to directly compete with the swinging arm for hybridization to the capture probe allowed us to estimate the effective local concentration of the swinging arm as ~250 µM for the 7-nm design and ~2.7 µM for the 14 nm construct ( Supplementary Figs. S27-28), demonstrating a steep distance dependence consistent with our predictions using a recent coarsegrained model 23 of the poly(T) 20 arm ( Supplementary Fig. S29 and Table S4).…”

Correlation Force Spectroscopy for Single Molecule Measurements

“…Some of the biological actions of ssDNA depend on its mechanical properties [51], which has motivated studies of the mechanical properties of ssDNA by single molecule force spectroscopy (SMFS) [134]. Two trends are obvious in the literature: (i) most studies have been on long ssDNA (n > 100, where n is the number of nucleotides) [12] whereas short ssDNA (n < 100) is encountered in nature during replication and in important technologies such as recombinant DNA technology [2], (ii) most experiments have examined homopolymeric ssDNA [51,[134][135][136] whereas most natural DNA is of necessity heteropolymeric to carry the genetic code.…”

“…Two trends are obvious in the literature: (i) most studies have been on long ssDNA (n > 100, where n is the number of nucleotides) [12] whereas short ssDNA (n < 100) is encountered in nature during replication and in important technologies such as recombinant DNA technology [2], (ii) most experiments have examined homopolymeric ssDNA [51,[134][135][136] whereas most natural DNA is of necessity heteropolymeric to carry the genetic code. Thus, the current work addresses the important area of short, heteropolymeric ssDNA.…”

“…Thus, the current work addresses the important area of short, heteropolymeric ssDNA. Most previous work on heteropolymeric ssDNA utilized spectroscopic techniques such as Förster resonance energy transfer (FRET) or scattering techniques small angle X-ray scattering (SAXS) [51,133,134,136].…”

“…For example, the high values of the friction coefficient of a molecule enhances its resistance to folding [61,66,72,134,[141][142][143]. The friction coefficient is the force divided by the velocity.…”