Di Capua E, Müller B
Institute of Cell Biology, ETH Hönggerberg, Zürich, Switzerland.
EMBO J. 1987 Aug;6(8):2493-8. doi: 10.1002/j.1460-2075.1987.tb02531.x.
recA protein coats DNA co-operatively to form filaments approximately 100 A thick, which in the presence of ATP, and more stably so in the presence of the non-hydrolyzable analog ATP gamma S, have a helical appearance with a deep cleft in the protein coat. This protein helix follows the DNA helix, to which it imparts a new helicity of 18.5 bp per turn of 97 A pitch. Here we test the accessibility of the DNA in the complex to modification by dimethylsulfate, and find that the complexed DNA is approximately 2-fold more reactive on the major groove side than it was in B-DNA (methylation of guanine N7), while it is protected approximately 2-fold on the minor groove side (methylation of adenine N3), suggesting that the protein coats the DNA along the minor groove. Furthermore, N3 of cytosine, a residue involved in base pairing, is found exposed in complexes with single strands as it is in naked single-stranded DNA, while it remains inaccessible in complexes with double strands, suggesting that the latter is not melted at this stage of the strand exchange reaction.
RecA蛋白协同包裹DNA形成厚度约为100埃的细丝，在ATP存在时，且在不可水解的类似物ATPγS存在时更稳定，这些细丝呈现螺旋外观，蛋白外壳中有一个深裂缝。这种蛋白螺旋沿着DNA螺旋，每97埃螺距每圈赋予DNA新的18.5个碱基对的螺旋度。在这里，我们测试了复合物中DNA对硫酸二甲酯修饰的可及性，发现复合DNA在大沟侧的反应性比B-DNA中大约高2倍（鸟嘌呤N7甲基化），而在小沟侧受到大约2倍的保护（腺嘌呤N3甲基化），这表明蛋白沿着小沟包裹DNA。此外，胞嘧啶的N3（参与碱基配对的一个残基）在与单链的复合物中如在裸露的单链DNA中一样暴露，而在与双链的复合物中仍不可及，这表明在链交换反应的这个阶段双链没有解链。
