Laboratory of Molecular Genetics and Laboratory of Structural Biology, National Institute of Environmental Health Sciences, NIH, DHHS, Research Triangle Park, NC 27709, USA.
DNA Repair (Amst). 2011 Aug 15;10(8):897-902. doi: 10.1016/j.dnarep.2011.05.009. Epub 2011 Jun 23.
The major replicative DNA polymerases of S. cerevisiae (Pols α, δ, and ɛ) incorporate substantial numbers of ribonucleotides into DNA during DNA synthesis. When these ribonucleotides are not removed in vivo, they reside in the template strand used for the next round of replication and could potentially reduce replication efficiency and fidelity. To examine if the presence of ribonucleotides in a DNA template impede DNA synthesis, we determined the efficiency with which Pols α, δ, and ɛ copy DNA templates containing a single ribonucleotide. All three polymerases can replicate past ribonucleotides. Relative to all-DNA templates, bypass of ribo-containing templates is slightly reduced, to extents that depend on the identity of the ribo and the sequence context in which it resides. Bypass efficiencies for Pols δ and ɛ were increased by increasing the dNTP concentrations to those induced by cellular stress, and in the case of Pol ɛ, by inactivating the 3'-exonuclease activity. Overall, ribonucleotide bypass efficiencies are comparable to, and usually exceed, those for the common oxidative stress-induced lesion 8-oxo-guanine.
酿酒酵母的主要复制 DNA 聚合酶（Pols α、δ 和 ɛ）在 DNA 合成过程中会将大量的核糖核苷酸掺入 DNA 中。如果这些核糖核苷酸在体内没有被去除，它们就会存在于下一轮复制所使用的模板链中，这可能会降低复制效率和保真度。为了研究 DNA 模板中核糖核苷酸的存在是否会阻碍 DNA 合成，我们测定了 Pols α、δ 和 ɛ 复制含有单个核糖核苷酸的 DNA 模板的效率。这三种聚合酶都可以复制核糖核苷酸。与全-DNA 模板相比，含有核糖核苷酸的模板的绕过效率略有降低，其程度取决于核糖的身份和它所处的序列上下文。通过将 dNTP 浓度增加到细胞应激诱导的浓度，以及在 Pol ɛ 的情况下通过失活 3'-外切酶活性，可以提高 Pols δ 和 ɛ 的绕过效率。总的来说，核糖核苷酸绕过效率与常见的氧化应激诱导损伤 8-氧鸟嘌呤相当，通常甚至超过后者。
