Sun W, Mao C, Liu F, Seeman N C
Department of Chemistry, New York University, New York, NY 10003, USA.
J Mol Biol. 1998 Sep 11;282(1):59-70. doi: 10.1006/jmbi.1998.1991.
The Holliday junction is a central intermediate in the process of genetic recombination. The position of its branch-point can relocate through an isomerization known as branch migration. This migration occurs because the branch-point is flanked by homologous symmetry. All attempts at modeling the kinetics of branch migration have relied on the assumption that branch migration minima are sequence-independent. We have tested that assumption here, using a competition assay based on symmetric immobile branched junctions; these are junctions that cannot undergo branch migration, despite the fact that they are flanked by homology. The assay used is predicated on the non-association of strands displaced in the assay; we have tested this assumption, and have performed our experiments under conditions where we know that it is true. We have measured the free energy of relocating a branched junction from a fixed non-homologous sequence to all possible dimeric symmetric sequences. We find that the assumption of sequence-independence is often valid, but that it is not universally true. We find that the flanking sequences can have a marked effect on the free energy measured, both for extensions of symmetry and for reversals of flanking nucleotides. We have varied the temperature in our experiments, and have derived both enthalpies and entropies for the different sequences. The entropies are largely unfavorable, whereas the enthalpies are largely favorable; regardless of the signs of these quantities, we see that this is another system where enthalpy-entropy compensation is operative.
霍利迪连接体是基因重组过程中的核心中间体。其分支点的位置可通过一种称为分支迁移的异构化作用重新定位。这种迁移的发生是因为分支点两侧具有同源对称性。所有对分支迁移动力学进行建模的尝试都依赖于这样一种假设，即分支迁移最小值与序列无关。我们在此使用基于对称固定分支连接体的竞争试验对该假设进行了测试；这些连接体尽管两侧具有同源性，但无法进行分支迁移。所使用的试验基于试验中被置换链不发生缔合的假设；我们对该假设进行了测试，并在已知该假设成立的条件下进行了实验。我们测量了将分支连接体从固定的非同源序列重新定位到所有可能的二聚体对称序列的自由能。我们发现序列无关的假设通常是有效的，但并非普遍成立。我们发现侧翼序列对所测量的自由能有显著影响，无论是对于对称性的延伸还是侧翼核苷酸的反转。我们在实验中改变了温度，并得出了不同序列的焓和熵。熵在很大程度上是不利的，而焓在很大程度上是有利的；无论这些量的符号如何，我们都看到这是另一个焓 - 熵补偿起作用的系统。
