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首页> 外文期刊>Journal of Molecular Biology >DNA-sequence asymmetry directs the alignment of recombination sites in the FLP synaptic complex.
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DNA-sequence asymmetry directs the alignment of recombination sites in the FLP synaptic complex.

机译：DNA序列不对称性指导FLP突触复合物中重组位点的对齐。

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摘要

The FLP recombinase promotes site-specific recombination in the 2 micrometer circle of Saccharomyces cerevisiae. FLP recognizes a 48 bp target site (FLP recombination target, or FRT) consisting of three 13 bp protein binding sites, or symmetry elements, flanking an 8 bp spacer region. Efficient recombination also occurs with DNA substrates that have minimal FRT sites, consisting only of the spacer and two surrounding 13 bp symmetry elements arranged in inverse orientation; thus, the wild-type spacer sequence is the main asymmetric feature of the minimal recombination site. FLP carries out recombination with many minimal target sites bearing symmetric or asymmetric mutant spacer sequences; however, the overall directionality of recombination defined in terms of inversion or excision of a DNA domain is determined by spacer-sequence asymmetry. In order to evaluate the potential influence of spacer-sequence asymmetry on structures formed during early steps in recombination, we used electron microscopy to investigate the structure of the FLP synaptic complex, which is the intermediate protein-DNA complex involved in site pairing and strand exchange. Using linear substrate DNAs that have minimal FRTs with wild-type spacer sequences, we find that 85 to 90% of the FLP synaptic complexes examined contain the two FRTs aligned in parallel. This strong preference for parallel site alignment stands in contrast with prevailing models for lambda integrase-class recombination systems, which postulate antiparallel site alignment, and results from biophysical studies on synthetic, immobile four-way DNA junctions. Our results show that the strong preference for parallel alignment can be attributed to conformational preferences of Holliday junctions present in the synaptosome. Copyright 1999 Academic Press.
机译：FLP重组酶促进酿酒酵母2微米圈内的位点特异性重组。 FLP识别一个48 bp的靶位点（FLP重组靶或FRT），该位点由三个13 bp的蛋白结合位点或对称元件组成，位于8 bp的间隔区旁。具有最小FRT位点的DNA底物也会发生有效重组，该底物仅由间隔子和两个反向排列的13 bp对称对称元件组成。因此，野生型间隔区序列是最小重组位点的主要不对称特征。 FLP与许多带有对称或不对称突变间隔区序列的最小目标位点进行重组;然而，根据间隔区序列的不对称性确定了根据DNA结构域的倒置或切除而定义的重组的整体方向性。为了评估间隔序列不对称对重组早期步骤中形成的结构的潜在影响，我们使用电子显微镜研究了FLP突触复合物的结构，该复合物是参与位点配对和链交换的中间蛋白质-DNA复合物。 。使用具有最小的FRT和野生型间隔序列的线性底物DNA，我们发现所检查的FLP突触复合物的85至90％包含两个平行排列的FRT。这种对平行位点比对的强烈偏好与lambda整合酶类重组系统的流行模型形成对比，后者假定反平行位点比对，并且是对合成的，固定的四向DNA连接进行生物物理研究的结果。我们的结果表明，对平行比对的强烈偏好可以归因于突触小体中存在的霍利迪连接的构象偏好。版权所有1999，学术出版社。

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