Single Molecule Studies of Sequence Dependence Elasticity in DNA

Abstract

DNA looping is a vital regulatory process in the cell to maintain proper function and viability. In protein-mediated looping, the DNA's sequence dictates both protein binding sites and the local mechanical properties of the DNA. These mechanical properties, notably its elasticity and intrinsic curvature, govern the ease at which loops can form. To probe the effects of sequence dependence on elasticity and the loop formation process, single molecule experiments were performed on short segments (∼150bp) of DNA with varying amounts of AT and GC content between the protein-binding operators. Tethered particle motion (TPM) microscopy was employed to observe protein-mediated DNA loop formation in this system, and an analytical model of the loop formation process was used to calculate the elasticity of the DNA from the observed loop formation rates. For comparison, axial constant-force optical tweezers were used to directly stretch the same DNA molecules mechanically to determine their persistence length as a measure for their elasticity. Our results indicate that the intraoperator sequence has a larger effect on elasticity in the loop formation experiments than in the stretching experiments, which we attribute to different elasticity regimes when the DNA is strongly bent as in a DNA loop, compared to the thermally induced small curvature fluctuations in stretched DNA.