Thermal energy suppresses mutational defects in DNA unwinding at a yeast replication origin.
Thermal energy suppresses mutational defects in DNA unwinding at a yeast replication origin.
复制标题
热能抑制酵母复制起点 DNA 解旋中的突变缺陷。
DOI：
10.1073/pnas.87.7.2486
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发表时间：
1990
影响因子：
11.1
通讯作者：
Kowalski,D
中科院分区：
文献类型：
--
作者：
Umek,RM;Kowalski,D
酵母复制起点含有DNA序列元件，其生物活性与对负超螺旋质粒中的单链特异性核酸酶的超敏性相关。通过使用质粒拓扑异构体的二维凝胶电泳，我们证明了化学稳定的起源解旋占核酸酶的超敏性，此外，增加的热能有利于稳定的起源解旋在体外。在活细胞中，增加的热能可以抑制提高解旋核酸酶超敏元件的自由能成本的起源突变。具体而言，与23 ℃相比，在30 ℃下生长的细胞中自主复制序列（ARS）介导的质粒复制中的突变缺陷不太严重。我们的研究结果表明，在核酸酶超敏感元件的DNA解旋的能量是生物学上重要的。我们称核酸酶超敏感序列的DNA解旋元件（DUE），并建议它作为酵母复制酶进入DNA螺旋的入口站点。
Yeast replication origins contain a DNA sequence element whose biological activity correlates with hypersensitivity to single-strand-specific nucleases in negatively supercoiled plasmids. By using two-dimensional gel electrophoresis of plasmid topoisomers, we demonstrate that thermodynamically stable origin unwinding accounts for the nuclease hypersensitivity and, furthermore, that increased thermal energy facilitates stable origin unwinding in vitro. In living cells, increased thermal energy can suppress origin mutations that raise the free-energy cost for unwinding the nuclease-hypersensitive element. Specifically, mutational defects in autonomously replicating sequence (ARS)-mediated plasmid replication are less severe in cells grown at 30 degrees C as compared to 23 degrees C. Our findings indicate that the energetics of DNA unwinding at the nuclease-hypersensitive element are biologically important. We call the nuclease-hypersensitive sequence the DNA unwinding element (DUE) and propose that it serves as the entry site for yeast replication enzymes into the DNA helix.