Rief M, Clausen-Schaumann H, Gaub H E
Lehrstuhl für Angewandte Physik, München, Germany.
Nat Struct Biol. 1999 Apr;6(4):346-9. doi: 10.1038/7582.
Atomic force microscope-based single-molecule force spectroscopy was employed to measure sequence-dependent mechanical properties of DNA by stretching individual DNA double strands attached between a gold surface and an AFM tip. We discovered that in lambda-phage DNA the previously reported B-S transition, where 'S' represents an overstretched conformation, at 65 pN is followed by a nonequilibrium melting transition at 150 pN. During this transition the DNA is split into single strands that fully recombine upon relaxation. The sequence dependence was investigated in comparative studies with poly(dG-dC) and poly(dA-dT) DNA. Both the B-S and the melting transition occur at significantly lower forces in poly(dA-dT) compared to poly(dG-dC). We made use of the melting transition to prepare single poly(dG-dC) and poly(dA-dT) DNA strands that upon relaxation reannealed into hairpins as a result of their self-complementary sequence. The unzipping of these hairpins directly revealed the base pair-unbinding forces for G-C to be 20 +/- 3 pN and for A-T to be 9 +/- 3 pN.
基于原子力显微镜的单分子力谱技术被用于通过拉伸附着在金表面和原子力显微镜探针之间的单条DNA双链来测量DNA的序列依赖性力学性质。我们发现，在λ噬菌体DNA中，先前报道的在65皮牛（pN）时发生的B-S转变（其中“S”代表过度拉伸构象）之后，在150皮牛时会发生非平衡解链转变。在这个转变过程中，DNA会分裂成单链，在松弛时又会完全重新结合。通过与聚（dG-dC）和聚（dA-dT）DNA的对比研究来考察序列依赖性。与聚（dG-dC）相比，聚（dA-dT）中的B-S转变和解链转变都在显著更低的力下发生。我们利用解链转变制备了单条聚（dG-dC）和聚（dA-dT）DNA链，由于它们的自互补序列，这些链在松弛时会重新退火形成发夹结构。这些发夹结构的解链直接揭示了G-C碱基对的解链力为20±3皮牛，A-T碱基对的解链力为9±3皮牛。
