The transition mechanism of DNA overstretching: a microscopic view using molecular dynamics

Abstract

The overstretching transition in torsionally unconstrained DNA is studied by means of atomistic molecular dynamics simulations. The free-energy profile as a function of the length of the molecule is determined through the umbrella sampling technique providing both a thermodynamic and a structural characterization of the transition pathway. The zero-force free-energy profile is monotonic but, in accordance with recent experimental evidence, becomes two-state at high forces. A number of experimental results are satisfactorily predicted: (i) the entropic and enthalpic contributions to the free-energy difference between the basic (B) state and the extended (S) state; (ii) the longitudinal extension of the transition state and (iii) the enthalpic contribution to the transition barrier. A structural explanation of the experimental finding that overstretching is a cooperative reaction characterized by elementary units of approximately 22 base pairs is found in the average distance between adenine/thymine-rich regions along the molecule. The overstretched DNA adopts a highly dynamical and structurally disordered double-stranded conformation which is characterized by residual base pairing, formation of non-native intra-strand hydrogen bonds and effective hydrophobic screening of apolar regions.

Keywords: DNA overstretching transition; molecular dynamics simulation of DNA overstretching; structural dynamics and energetics of DNA overstretching; structural model of DNA overstretching.

Figure 1.

Force–extension relation of ds λ-DNA for a molecule that does not show hysteresis upon relaxation (temperature 25°C, 150 mM NaCl). Adapted from fig. 1b′ of ref. [13].

Figure 2.

Averaged conformations corresponding to an average distance between consecutive base pairs of (from bottom to top) 0.34, 0.40, 0.46, 0.52 and 0.58 nm. AT base pairs are represented in yellow; CG base pairs in green.

Figure 3.

(a) Decrease in the percentage of native base stackings (squares) and in the global percentage of native hydrogen bonds (circles) in relation to increase in the average distance between consecutive base pairs. (b) Average base pair distance in the AT-rich region (diamonds) and in the CG-rich region (triangles).

Figure 4.

Enlarged image for the conformation with base pair distance of 0.58 nm. Carbons of the AT base pairs are yellow; those of CG base pairs are green. Nitrogens are blue, phosphates orange, oxygens light red and hydrogens white. Hydrogen bonds are shown by cyan lines. Circles identify the binding of bases to the phosphate backbone of the opposite strand. Arrows show nucleoside stacking to other nucleosides belonging to the opposite strand (yellow for the AT base pair, green for the CG base pair).

Figure 5.

Comparison of the hydrophobicity surface between the basic (b) and the overstretched (a) conformation. Red corresponds to highly hydrophilic atoms and blue to strongly hydrophobic ones. White is neutrality. Phosphates are orange. The right portion of the overstretched conformation corresponds to the CG-rich region of the molecule that retains the native base pairing. The left portion is the AT-rich one. In spite of their inherent structural differences at the hydrogen bonding level, both regions show the same large-scale hydrophobic organization: the negative backbones are exposed to water and are kept separated from each other by the hydrophobic nucleosides.

Figure 6.

Energy profiles for the overstretching of the 23-mer. (a) Free-energy profile, computed by means of an umbrella sampling analysis over 2 ns trajectories. In the inset, the free-energy profile at the coexistence force F_e. (b) Enthalpic (ΔE) and entropic (TΔS) contributions to the free-energy profile. In the inset, the van der Waals contribution (ΔE_vdw) to the free-energy profile. The dashed lines are two independent linear fits to the data above and below x_bp = 0.46 nm (identified by the arrow).