Replication of eukaryotic genomes relies on the family B DNA polymerases Pol α, Pol δ, and Pol ε. All of these enzymes coordinate an iron-sulfur (FeS) cluster, but the function of this cofactor has remained largely unclear. Here, we show that the FeS cluster in the catalytic subunit of human Pol δ is coordinated by four invariant cysteines of the C-terminal CysB motif. FeS cluster loss causes a partial destabilisation of the four-subunit enzyme, a defect in double-stranded DNA binding, and compromised polymerase and exonuclease activities. Importantly, complex stability, DNA binding, and enzymatic activities are restored in the presence of proliferating cell nuclear antigen. We further show that also more subtle changes to the FeS cluster-binding pocket that do not abolish FeS cluster binding can have repercussions on the distant exonuclease domain and render the enzyme error prone. Our data hence suggest that the FeS cluster in human Pol δ is an important co-factor that despite its C-terminal location has an impact on both DNA polymerase and exonuclease activities, and can influence the fidelity of DNA synthesis.

中文翻译：

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人类DNA聚合酶δ需要铁硫簇才能实现高保真DNA合成。

真核基因组的复制依赖于B族DNA聚合酶Polα，Polδ和Polε。所有这些酶均能协调铁硫（FeS）簇，但该辅因子的功能仍不清楚。在这里，我们显示人类Polδ催化亚基中的FeS簇由C端CysB图案的四个不变半胱氨酸协调。FeS簇丢失会导致四亚基酶部分不稳定，双链DNA结合缺陷以及聚合酶和核酸外切酶活性受损。重要的是，在增殖细胞核抗原的存在下，可以恢复复杂的稳定性，DNA结合和酶促活性。我们进一步表明，对FeS簇结合口袋的更微妙的变化（不消除FeS簇结合）可以对遥远的核酸外切酶结构域产生影响，并使酶容易出错。因此，我们的数据表明，人Polδ中的FeS簇是重要的辅助因子，尽管其C端位置对DNA聚合酶和核酸外切酶活性均具有影响，并且可以影响DNA合成的保真度。