Barone Flavia, McCulloch Scott D, Macpherson Peter, Maga Giovanni, Yamada Masami, Nohmi Takehiko, Minoprio Anna, Mazzei Filomena, Kunkel Thomas A, Karran Peter, Bignami Margherita
Unit of Experimental Carcinogenesis, Department of Environment and Primary Prevention, Istituto Superiore di Sanità, Viale Regina Elena 299, 00161 Rome, Italy.
DNA Repair (Amst). 2007 Mar 1;6(3):355-66. doi: 10.1016/j.dnarep.2006.11.002. Epub 2006 Dec 26.
2-Hydroxyadenine (2-OH-A), a product of DNA oxidation, is a potential source of mutations. We investigated how representative DNA polymerases from the A, B and Y families dealt with 2-OH-A in primer extension experiments. A template 2-OH-A reduced the rate of incorporation by DNA polymerase alpha (Pol alpha) and Klenow fragment (Kf(exo-)). Two Y family DNA polymerases, human polymerase eta (Pol eta) and the archeal Dpo4 polymerase were affected differently. Bypass by Pol eta was very inefficient whereas Dpo4 efficiently replicated 2-OH-A. Replication of a template 2-OH-A by both enzymes was mutagenic and caused base substitutions. Dpo4 additionally introduced single base deletions. Thermodynamic analysis showed that 2-OH-A forms stable base pairs with T, C and G, and to a lesser extent with A. Oligonucleotides containing 2-OH-A base pairs, including the preferred 2-OH-A:T, were recognized by the human MutSalpha mismatch repair (MMR). MutSalpha also recognized 2-OH-A located in a repeat sequence that mimics a frameshift intermediate.
2-羟基腺嘌呤（2-OH-A）是DNA氧化的产物，是潜在的突变源。我们在引物延伸实验中研究了来自A、B和Y家族的代表性DNA聚合酶如何处理2-OH-A。模板2-OH-A降低了DNA聚合酶α（Polα）和克列诺片段（Kf(exo-)）的掺入速率。两种Y家族DNA聚合酶，人聚合酶η（Polη）和古细菌Dpo4聚合酶受到的影响不同。Polη的跨损伤合成效率非常低，而Dpo4能有效复制2-OH-A。两种酶对模板2-OH-A的复制都具有致突变性并导致碱基替换。Dpo4还会引入单碱基缺失。热力学分析表明，2-OH-A与T、C和G形成稳定的碱基对，与A形成的碱基对程度较小。包含2-OH-A碱基对（包括优选的2-OH-A:T）的寡核苷酸可被人MutSα错配修复（MMR）识别。MutSα还能识别位于模拟移码中间体的重复序列中的2-OH-A。
