Strand displacement during deoxyribonucleic acid synthesis at single strand breaks.
Strand displacement during deoxyribonucleic acid synthesis at single strand breaks.
复制标题
脱氧核糖核酸合成过程中单链断裂处的链置换。
DOI：
10.1016/s0021-9258(18)62341-5
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发表时间：
1971
期刊：
影响因子：
--
通讯作者：
C. Richardson
中科院分区：
文献类型：
--
作者：
Y. Masamune;C. Richardson
利用含有单磷酸二酯键中断（缺口）的环状双链DNA模板，研究了DNA聚合酶合成DNA的方法。大肠杆菌dna聚合酶可以通过核苷酸共价附着在引物链的3'端而在缺口处启动合成。合成的初始阶段伴随着e的5‘水解活性从缺口的5’端去除核苷酸。共聚合酶（nick翻译）。在加入10到50个核苷酸后，引物链的5‘端被引物链的3’端所取代。消除theE的5'-水解活性。聚合酶不能阻止这种链移位。相反，噬菌体T4聚合酶不能在切口处启动合成。在这些研究中，多核苷酸连接酶已被用于识别刻痕和测量它们在链位移过程中的消失。在链置换阶段合成的DNA对e的外切酶I和III都很敏感。杆菌。这一观察结果表明，在新合成的DNA的单链和双链形式之间存在一种平衡。第三阶段更广泛的合成产生的DNA产物，即使在变性后也能抵抗外切酶I，表明其结构是自互补的。在含有一个缺口的环状PM2 DNA上合成的这种产物的电子显微照片显示，从完整的圆形延伸出一个分支。随后的合成产生了多个分支。
Synthesis of DNA by DNA polymerases has been studied by using circular duplex DNA templates which contain single phosphodiester bond interruptions (nicks).Escherichia coliDNA polymerase can initiate synthesis at nicks by the covalent attachment of nucleotides to the 3' end of the primer strand. An initial phase of synthesis is accompanied by the removal of nucleotides from the 5' end of the nick by the 5'-hydrolytic activity of theE. colipolymerase (nick translation). After the incorporation of 10 to 50 nucleotides, the 5' end of the primer strand is displaced by the growing 3' end of the primer strand. Elimination of the 5'-hydrolytic activity of theE. colipolymerase does not prevent this strand displacement. In contrast, phage T4 polymerase cannot initiate synthesis at nicks. In these studies polynucleotide ligase has been used to identify nicks and to measure their disappearance during strand displacement.The DNA synthesized during the phase of strand displacement is sensitive to both exonucleases I and III ofE. coli. This observation suggests that an equilibrium exists between single and double stranded forms of the newly synthesized DNA. A third phase of more extensive synthesis yields a DNA product which is resistant to exonuclease I even after denaturation, suggesting a self-complementary structure. Electron micrographs of such a product synthesized on circular PM2 DNA containing one nick per molecule reveal a single branch extending from the intact circle. Subsequent synthesis gives rise to multiple branches.