Double-chain model. (a) 3D representation of the interwound bead chains. The particles comprising the duplex unit are labeled. (b) 2D schematic. Complementary beads (e.g., and ) touch i.e., are separated by twice the bead radius at mechanical equilibrium. The intrachain separation of particles and is labeled as . In the actual 3D structure, noncomplementary interchain beads (e.g., and ) approximately touch at equilibrium.
Trajectory snapshots. The white regions indicate content, gray regions content (base pairs 43-51 in this 141-bp structure), and the region labeled “” locations undergoing rotation at constant angular velocity. (a) : ringlike initial conformation. (b) : increasingly negative superhelicity resulting from continual rotation of both ends of the structure has produced left-handed, plectonemic interwinding. (c) : the region, which has localized to the apex of a supercoil, has opened. (d) : additional duplex opening has occurred in regions adjacent to the initial site of strand separation, unwinding right-handed twist, and relaxing left-handed supercoils.
Comparison of melting profiles with transition probabilities. (a), (c), (e), and (g) plot the time in milliseconds vs base pair position. The black areas indicate regions of the model structure that have irreversibly melted, and white areas indicate regions that are unmelted. The gray bar demarcates the 3-bp site of initial melting. (b), (d), (f), and (h) plot the probability of duplex opening (at superhelix density ) vs base pair position, calculated using WEBSIDD (see discussion). The gray bar again demarcates the site of initial melting. (a) and (b) correspond to a 141 base pair sequence in which base pairs 43-51 are of content, and the remaining base pairs are of content. In the sequence represented by (c) and (d), the region spans base pairs 67-75, and in the sequence represented by (e) and (f) this region spans base pairs 94-102. The sequence represented by (g) and (h) is entirely of content. For all sequences containing an region, the site of initial strand separation coincides with the transition probability profile predicted by WEBSIDD. Comparison of (g) and (h) suggests that, in the case of all- content, factors other than the base sequence, such as supercoil geometry, determine the location of the initial strand opening. The time of onset of melting in all dynamic simulations is after is reached.
Localization of denatured regions to loops. (a) [cf. Fig. 2(c)] is a snapshot taken from the simulation corresponding to the melting profile of Fig. 3(a) near the time of initial melting. Similarly, (b) corresponds to Fig. 3(c), (c) to Fig. 3(e), and (d) to Fig. 3(g). sites are the first to open in structures in which they are present. In (d), the entire structure is of content. In all simulations, initial duplex opening occurs in the terminal loop of a supercoil, suggesting both that the reduced effective bending stiffness associated with regions organizes supercoil geometry and that this geometry influences the site of initial opening.
Time dependence of topological and geometrical properties. Calculations of the linking number , writhe , twist , and superhelix density vs time (in milliseconds) correspond to the simulation represented by Figs. 3(c) and 4(b). At early times, continually decreasing is partitioned nearly exclusively to , and the initially planar duplex axis maintains . At just after 0.1 ms, a mechanical threshold is reached, after which becomes increasingly negative (reaching a maximum value near ) as the structure plectonemically supercoils, while maintains an approximately constant average value. Then, at approximately 0.3 ms [cf. Fig. 3(c)], localized strand separation initiates, leading to the unwinding of about five turns and a relaxation to by 0.5 ms. Over the course of the simulation, a value of the superhelix density is reached.
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