Shimizu K, Santocanale C, Ropp P A, Longhese M P, Plevani P, Lucchini G, Sugino A
Laboratory of Molecular Genetics, National Institute of Environmental Health Sciences, National Institutes of Health, Research Triangle Park, North Carolina 27709.
J Biol Chem. 1993 Dec 25;268(36):27148-53.
A new DNA polymerase activity was identified and purified to near homogeneity from extracts of mitotic and meiotic cells of the yeast Saccharomyces cerevisiae. This activity increased at least 5-fold during meiosis, and it was shown to be associated with a 68-kDa polypeptide as determined by SDS-polyacrylamide gel electrophoresis. This new DNA polymerase did not have any detectable 3'-->5' exonuclease activity and preferred small gapped DNA as a template-primer. The activity was inhibited by dideoxyribonucleoside 5'-triphosphates and N-ethylmaleimide but not by concentrations of aphidicolin which completely inhibit either DNA polymerases I (alpha), II (epsilon), or III (delta). Since no polypeptide(s) in the extensively purified DNA polymerase fractions cross-reacted with antibodies raised against yeast DNA polymerases I, II, and III, we called this enzyme DNA polymerase IV. The DNA polymerase IV activity increased at least 10-fold in a yeast strain overexpressing the gene product predicted from the YCR14C open-reading frame (identified on S. cerevisiae chromosome III and provisionally called POLX), while no activity was detected in a strain where POLX was deleted. These results strongly suggest that DNA polymerase IV is encoded by the POLX gene and is a probable homolog of mammalian DNA polymerase beta.
从酿酒酵母有丝分裂和减数分裂细胞提取物中鉴定并纯化出一种新的DNA聚合酶活性，其纯度接近均一。该活性在减数分裂期间至少增加了5倍，通过SDS-聚丙烯酰胺凝胶电泳测定，它与一种68 kDa的多肽相关。这种新的DNA聚合酶没有任何可检测到的3'→5'核酸外切酶活性，并且更喜欢小缺口DNA作为模板引物。该活性受到双脱氧核糖核苷5'-三磷酸和N-乙基马来酰亚胺的抑制，但不受完全抑制DNA聚合酶I（α）、II（ε）或III（δ）的浓度的阿非科林的抑制。由于在广泛纯化的DNA聚合酶组分中没有多肽与针对酵母DNA聚合酶I、II和III产生的抗体发生交叉反应，我们将这种酶称为DNA聚合酶IV。在过表达由YCR14C开放阅读框（在酿酒酵母染色体III上鉴定并暂称为POLX）预测的基因产物的酵母菌株中，DNA聚合酶IV活性至少增加了10倍，而在删除POLX的菌株中未检测到活性。这些结果强烈表明，DNA聚合酶IV由POLX基因编码，并且可能是哺乳动物DNA聚合酶β的同源物。
