DNA DOUBLE-CROSSOVER MOLECULES
DNA DOUBLE-CROSSOVER MOLECULES
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DOI：
10.1021/bi00064a003
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发表时间：
1993-04-06
期刊：
影响因子：
2.9
通讯作者：
SEEMAN, NC
中科院分区：
文献类型：
--
作者：
FU, TJ;SEEMAN, NC
螺旋结构域之间含有两个交叉位点的 DNA 分子被认为是涉及双链断裂的重组过程中的中间体。我们在寡核苷酸系统中模拟了这些双交叉结构。尽管在设计这些分子时必须指定螺旋结构域的相对方向，但分子有两大类：平行分子 DP 和反平行分子 DA。交叉点之间的距离必须指定为半匝的倍数，以避免系统中出现扭转应力；因此，还有两个进一步的细分，即由奇数 O 和偶数 E 半匝数分隔的双交叉分子。此外，交叉点之间半匝数为奇数的平行分子必须分为具有过量主槽或宽槽分离的分子，W，或具有过量小槽或窄槽分离的分子，N。我们已经构建了所有这五类的模型：DAE、DAO、DPE、DPOW 和 DPON。已构建出在交叉点之间含有 1 和 2 个螺旋圈的 DPE 分子； DAE 分子在交叉之间包含 1 个螺旋圈，DAO、DPOW 和 DPON 分子在交叉之间包含 1.5 个螺旋圈。没有一个平行分子的行为是良好的；当在天然聚丙烯酰胺凝胶上观察时，这些分子要么解离，要么形成多聚体。相反，当以这种方式测定时，反平行分子形成单条带。这些分子的羟基自由基自动足迹分析揭示了在预期交叉和闭塞位点的保护，表明所有复合物都包含在交叉之间大致共面的线性螺旋轴。然而，交叉点之间有 2 圈的 DPOW 分子和 DPE 分子在交叉点之间的部分显示出保护性降低，这表明它们的螺旋可能会因电荷排斥而弯曲。我们得出的结论是，如果平行双交叉之间的螺旋要参与重组，则它们必须彼此屏蔽或扭曲线性。我们分析了双交叉分子中支链迁移和交叉异构化的可能性。平行分子除了同源性之外不需要序列对称性来进行分支迁移，但是反平行分子的序列对称性要求将迁移限制为直接重复的片段，这些片段迭代交叉之间的序列。交叉异构化在平行双交叉分子中似乎是一个非常复杂的过程，这表明如果它发生在细胞内，它可能是由拓扑异构酶催化的。
DNA molecules containing two crossover sites between helical domains have been suggested as intermediates in recombination processes involving double-strand breaks. We have modeled these double-crossover structures in an oligonucleotide system. Whereas the relative orientations of the helical domains must be specified in designing these molecules, there are two broad classes of the molecules, the parallel, DP, and antiparallel, DA, molecules. The distance between crossover points must be specified as multiples of half-turns, in order to avoid torsional stress in this system; hence, there are two further subdivisions, those double-crossover molecules separated by odd, O, and even, E, numbers of half-turns. In addition, the parallel molecules with odd numbers of half-turns between crossovers must be divided into those with an excess major or wide-groove separation, W, or those with an excess minor- or narrow-groove separation, N. We have constructed models of all five of these classes, DAE, DAO, DPE, DPOW, and DPON. DPE molecules containing 1 and 2 helical turns between crossovers have been constructed; the DAE molecule contains 1 turn between crossovers, and the DAO, DPOW, and DPON molecules contain 1.5 helical turns between crossovers. None of the parallel molecules is well-behaved; the molecules either dissociate or form multimers when visualized on native polyacrylamide gels. In contrast, antiparallel molecules form single bands when assayed in this fashion. Hydroxyl radical autofootprinting analysis of these molecules reveals protection at expected sites of crossover and of occlusion, suggesting that all the complexes contain linear helix axes that are roughly coplanar between crossovers. However, the DPOW molecule and the DPE molecule with 2 turns between crossovers show decreased protection in the portion between crossovers, suggesting that their helices may bow in response to charge repulsion. We conclude that the helices between parallel double crossovers must be shielded from each other or distorted from linearity if they are to participate in recombination. We have analyzed the possibilities of branch migration and crossover isomerization in double-crossover molecules. Parallel molecules need no sequence symmetry beyond homology to branch migrate, but the sequence symmetry requirements for antiparallel molecules restrict migration to directly repetitive segments that iterate the sequence between crossovers. Crossover isomerization appears to be a very complex process in parallel double-crossover molecules, suggesting that it may be catalyzed by topoisomerases if it occurs within the cell.