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he mechanical properties of DNA molecules are important to many biological behaviors, such as DNA binding to proteins, packaging in bacteriophage capsids, and wrapping around nucleosomes. Experimental results have shown that when B-DNA is overstretched, it
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transforms into S-DNA which is about 1.7 times the contour length of B-DNA (this is known as overstretching transition). Moreover, since DNA molecules have negatively charged phosphate groups along the double helix, the overstretching transition from B-DNA to S-DNA is sensitive to surrounding solution conditions.
Using optical tweezers, the ionic effects of NaCl solution on the overstretching transition of B-DNA were experimentally studied. The manipulation to stretch a single B-form�-DNA molecule is shown in Figure 1. As the diameter of a single DNA molecule is only about 2 nm, this is invisible even in the view of 100x magnification oil immersed objective lens and difficult to be trapped directly by laser beam. Instead of the DNA molecule, two streptavidin coated polystyrene microspheres were trapped in the experiment. These two microspheres were specifically bound to the two ends of the biotin labeled DNA molecule. One of the microspheres was attached to the bottom of the sample chamber and the other was captured and held stationary by the laser trap. By moving the sample chamber via the microscope stage, the DNA molecule was stretched until the trapped bead was detached from the laser trap. By varying the laser power, different stretching forces could be applied. The absolute extension of the molecule was determined by measuring the distance between the centers of the two microspheres using an image captured by a CCD camera. Force-extension measurements for stretching a single DNA molecule were then carried out.
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A three-dimensional model has been proposed to explain the experimental results. In this modified analytical model, bending deformation of DNA backbones, cooperativity of base-stacking interactions, electrostatic interactions, and spatial effects of DNA double helix structure are all taken into account. The key contribution of this model is that the electrostatic energy is explicitly given as a function of folding angle and salt concentration. This model also introduces a new parameter to account for the cooperativity of base-stacking interactions. The Metropolis Monte Carlo method was applied to simulate this model. As shown in Figure 2, numerical results obtained from the modified model are consistent with those from experimental observations. The results show that when the sodium ion concentration increases from 0.909 mM to 909 mM, the overstretching transition force increases from 43.01 pN to 66.02 pN. The overstretching transition force is linearly related to the natural logarithm of sodium ion concentration. This relation suggests that the overstretching transition force is sensitive to the concentration level of sodium salt solutions at low ion concentrations.
This work was done in collaboration with Assoc Prof WT Liu, Assoc Prof CT Lim, Dr H Chen & Ms HX Fu (formerly with the CE Dept).
Contact person
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Prof CG Koh
Tel: 65162163,
Fax: 67791635
E-mail: cgkoh@nus.edu.sg
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