DNA polymerase delta isolated from Schizosaccharomyces pombe contains five subunits
DNA polymerase delta isolated from Schizosaccharomyces pombe contains five subunits
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DOI：
10.1073/pnas.94.21.11244
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发表时间：
1997-10-14
影响因子：
11.1
通讯作者：
Hurwitz, J
中科院分区：
文献类型：
--
作者：
Zuo, SJ;Gibbs, E;Hurwitz, J
DNA 聚合酶 delta (pol delta) 在 DNA 复制、修复和重组中发挥着重要作用。我们从粟酒裂殖酵母中纯化了 pol δ 超过 10(3) 倍，并证明纯化的粟酒裂殖酵母 pol δ 的聚合酶活性完全依赖于增殖细胞核抗原和复制因子 C。纯化级分的 SDS/PAGE 分析表明 pol δ 复合物由五个亚基组成，这些亚基以 125、55、54、42 和 22 kDa 的表观分子量迁移。 Western blot 分析表明，125、55 和 54 kDa 的蛋白分别是大催化亚基 (Pol3)、Cdc1 和 Cdc37。其他两个亚基 p42 和 p22 的身份是在对所得肽进行蛋白水解消化和序列分析后确定的。源自 p22 亚基的肽序列表明，该亚基与 Cdm1 相同，Cdm1 先前被鉴定为温度敏感的 cdc1-P13 突变体的多拷贝抑制子，而源自 p42 亚基的肽序列与先前未表征的位于粟酒裂殖酵母染色体 1 上的 ORF 相同。
DNA polymerase delta (pol delta) plays an essential role in DNA replication, repair, and recombination. We have purified pol delta from Schizosaccharomyces pombe more than 10(3)-fold and demonstrated that the polymerase activity of purified S. pombe pol delta is completely dependent on proliferating cell nuclear antigen and replication factor C. SDS/PAGE analysis of the purified fraction indicated that the pol delta complex consists of Five subunits that migrate with apparent molecular masses of 125, 55, 54, 42, and 22 kDa. Western blot analysis indicated that the 125, 55, and 54 kDa proteins are the large catalytic subunit (Pol3), Cdc1, and Cdc37, respectively. The identity of the other two subunits, p42 and p22, was determined following proteolytic digestion and sequence analysis of the resulting peptides. The peptide sequences derived from the p22 subunit indicated that this subunit is identical to Cdm1, previously identified as a multicopy suppressor of the temperature-sensitive cdc1-P13 mutant, whereas peptide sequences derived from the p42 subunit were identical to a previously uncharacterized ORF located on S. pombe chromosome 1.