Zakharova Svetlana S, Jesse Wim, Backendorf Claude, Egelhaaf Stefan U, Lapp Alain, van der Maarel Johan R C
Leiden Institute of Chemistry, Leiden University, 2300 RA Leiden, The Netherlands.
Biophys J. 2002 Aug;83(2):1106-18. doi: 10.1016/S0006-3495(02)75234-X.
With a view to determine the configuration and regularity of plectonemically supercoiled DNA, we have measured the small angle neutron scattering from pUC18 plasmid in saline solutions. Furthermore, we have derived the mathematical expression for the single chain scattering function (form factor) of a superhelical structure, including the longitudinal and transverse interference over the plectonemic pitch and radius, respectively. It was found that an interwound configuration describes the data well, provided interactions among supercoils are accounted for in the second virial approximation. The opening angle was observed to be relatively constant and close to 58 degrees, but it was necessary to include a significant distribution in radius and pitch. For diluted supercoils with vanishing mutual interaction, the derived structural results agree with independent measurements, including the distribution in linking number deficit as determined by gel electrophoresis. With increasing plasmid concentration, prior and covering the transition to the liquid-crystalline phase, the radius and pitch are seen to decrease significantly. The latter observation shows that compaction of negatively supercoiled DNA by confinement results in a decrease in writhing number at the cost of a positive twist exerted on the DNA duplex. It is our conjecture that the free energy associated with this excess twist is of paramount importance in controlling the critical boundaries pertaining to the transition to the anisotropic, liquid-crystalline phase.
为了确定麻花状超螺旋DNA的结构和规律,我们测量了盐溶液中pUC18质粒的小角中子散射。此外,我们推导了超螺旋结构单链散射函数(形状因子)的数学表达式,分别包括在麻花状螺距和半径上的纵向和横向干涉。结果发现,只要在第二维里近似中考虑超螺旋之间的相互作用,缠绕结构就能很好地描述数据。观察到开口角相对恒定,接近58度,但有必要在半径和螺距上纳入显著的分布。对于相互作用消失的稀释超螺旋,推导得到的结构结果与独立测量结果一致,包括凝胶电泳测定的连接数亏缺分布。随着质粒浓度的增加,在进入液晶相转变之前和过程中,可以看到半径和螺距显著减小。后一观察结果表明,通过限制对负超螺旋DNA进行压缩会导致扭曲数减少,代价是在DNA双链体上施加正扭转。我们推测,与这种过量扭转相关的自由能在控制与向各向异性液晶相转变相关的临界边界方面至关重要。