Morfill Julia, Kühner Ferdinand, Blank Kerstin, Lugmaier Robert A, Sedlmair Julia, Gaub Hermann E
Lehrstuhl für Angewandte Physik & Center for Nanoscience, Ludwig-Maximilians-Universität München, Munich, Germany.
Biophys J. 2007 Oct 1;93(7):2400-9. doi: 10.1529/biophysj.107.106112. Epub 2007 Jun 8.
Stretching experiments with long double-stranded DNA molecules in physiological ambient revealed a force-induced transition at a force of 65 pN. During this transition between B-DNA and highly overstretched S-DNA the DNA lengthens by a factor of 1.7 of its B-form contour length. Here, we report the occurrence of this so-called B-S transition in short duplexes consisting of 30 basepairs. We employed atomic-force-microscope-based single molecule force spectroscopy to explore the unbinding mechanism of two short duplexes containing 30 or 20 basepairs by pulling at the opposite 5' termini. For a 30-basepair-long DNA duplex the B-S transition is expected to cause a length increase of 6.3 nm and should therefore be detectable. Indeed 30% of the measured force-extension curves exhibit a region of constant force (plateau) at 65 pN, which corresponds to the B-S transition. The observed plateaus show a length between 3 and 7 nm. This plateau length distribution indicates that the dissociation of a 30-basepair duplex mainly occurs during the B-S transition. In contrast, the measured force-extension curves for a 20-basepair DNA duplex exhibited rupture forces below 65 pN and did not show any evidence of a B-S transition.
在生理环境中对长双链DNA分子进行的拉伸实验显示,在65皮牛的力作用下会发生力诱导转变。在B-DNA和高度过度拉伸的S-DNA之间的这种转变过程中,DNA长度增加到其B型轮廓长度的1.7倍。在此,我们报告了由30个碱基对组成的短双链体中这种所谓的B-S转变的发生情况。我们采用基于原子力显微镜的单分子力谱技术,通过在相对的5'末端进行拉伸,来探究两个分别包含30个或20个碱基对的短双链体的解链机制。对于一个30个碱基对长的DNA双链体,B-S转变预计会导致长度增加6.3纳米,因此应该是可检测到的。实际上,30%的测量力-伸长曲线在65皮牛处呈现出一个恒力区域(平台期),这对应于B-S转变。观察到的平台期长度在3到7纳米之间。这种平台期长度分布表明,一个30个碱基对双链体的解离主要发生在B-S转变期间。相比之下,一个20个碱基对DNA双链体的测量力-伸长曲线显示断裂力低于65皮牛,并且没有显示出任何B-S转变的迹象。