Albany 2013: Book of Abstracts
June 11-15 2013
©Adenine Press (2012)
Kinetics of DNA overstretching: Melting vs B-to-S transition
Single B-form DNA molecules undergo an overstretching transition at force Fov to a ~1.7-fold longer form when stretched. The nature of overstretched DNA has been debated for over 10 years. Either peeled (PL DNA), internally melted (M DNA), or unwound double-helical (S DNA) forms of overstretched DNA have been suggested. Here we characterize the kinetics of the overstretching transition in polymeric torsionally unconstrained double stranded (ds) DNA molecules. We pull ~50 Kbp λ−DNA molecules using optical tweezers with rates ν ~10 nm/s to 5.104 nm/s, (over-stretching time between 0.2 and 103 s). The Fov (ν, [Na+]) dependence measured over a broad range of rates and solution ionic strength suggests the existence of all three forms of the overstretched DNA. Thus, at [Na+]>50mM and the stretching time >>1s, internal melting dominates overstretching. This B-to-M transition is highly cooperative (involves ~100 bp), and slow (on/off time ~1000s). Faster overstretching during ≤1 s leads to B-to-S DNA transition, which is less cooperative (involves ~10 bp) and faster (on/off time ~1s). In contrast, in lower salt ([Na+]<50mM) the overstretching during >1s leads to DNA peeling. However, on the faster time scale of 0.2-1 s, even in low salt, the DNA overstretches into S DNA, as peeling becomes kinetically prohibited. Our conclusions are supported by several independent lines of evidence, including the salt and rate dependence of both the slope of the overstretched DNA force-extension curve and the value of the second transition force (from M or PL DNA into S DNA).
Micah J. McCauley 1