The structure and polymerase-recognition mechanism of the crucial adaptor protein AND-1 in the human replisome
The structure and polymerase-recognition mechanism of the crucial adaptor protein AND-1 in the human replisome
复制标题
人类复制体中关键接头蛋白AND-1的结构和聚合酶识别机制
DOI：
10.1074/jbc.m116.758524
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发表时间：
2017-06-09
影响因子：
4.8
通讯作者：
Liang, Huanhuan
中科院分区：
文献类型：
--
作者：
Guan, Chengcheng;Li, Jun;Liang, Huanhuan
真核细胞中的DNA复制是由一种称为复制体的多蛋白复合物完成的，该复合体由解旋酶、聚合酶和接头分子组成。人酸性核质DNA结合蛋白1 (and -1)，又称WD重复和高迁移率组（HMG）盒DNA结合蛋白1 (WDHD1)，是DNA复制的重要衔接分子。虽然关于AND-1酵母同源物的结构信息是可用的，但是关于人类AND-1蛋白如何将滞后链DNA聚合酶α (pol α)锚定在DNA解旋酶复合体（Cdc45-MCM2-7-GINS， CMG）上的机制细节还有待阐明。本文报道了人类and -1基因n端WD40和SepB结构域的结构，以及c端HMG结构域的生化分析。我们发现AND-1以SepB结构域介导的三聚体形式存在。突变体研究结果表明，SepB结构域中的一个带正电的凹槽为pol α提供了结合位点。与其在出芽酵母中的同源蛋白不同，人的AND-1被招募到CMG复合体中，由除Go Ichi Ni San之外的未知参与者介导。此外，我们发现AND-1在体外通过其c端HMG结构域与DNA结合。总之，我们的发现为人类AND-1功能的机制细节提供了重要的见解，促进了我们对真核生物复制过程中复制体形成的理解。
DNA replication in eukaryotic cells is performed by a multiprotein complex called the replisome, which consists of helicases, polymerases, and adaptor molecules. Human acidic nucleoplasmic DNA-binding protein 1 (AND-1), also known as WD repeat and high mobility group (HMG)-box DNA-binding protein 1 (WDHD1), is an adaptor molecule crucial for DNA replication. Although structural information for the AND-1 yeast ortholog is available, the mechanistic details for how human AND-1 protein anchors the lagging-strand DNA polymerase α (pol α) to the DNA helicase complex (Cdc45-MCM2–7-GINS, CMG) await elucidation. Here, we report the structures of the N-terminal WD40 and SepB domains of human AND-1, as well as a biochemical analysis of the C-terminal HMG domain. We show that AND-1 exists as a homotrimer mediated by the SepB domain. Mutant study results suggested that a positively charged groove within the SepB domain provides binding sites for pol α. Different from its ortholog protein in budding yeast, human AND-1 is recruited to the CMG complex, mediated by unknown participants other than Go Ichi Ni San. In addition, we show that AND-1 binds to DNA in vitro, using its C-terminal HMG domain. In conclusion, our findings provide important insights into the mechanistic details of human AND-1 function, advancing our understanding of replisome formation during eukaryotic replication.