The C-terminal domain of yeast PCNA is required for physical and functional interactions with Cdc9 DNA ligase.
The C-terminal domain of yeast PCNA is required for physical and functional interactions with Cdc9 DNA ligase.
复制标题
酵母菌PCNA的C末端结构域是与Cdc9 DNA连接酶的物理和功能相互作用所必需的。
DOI：
10.1093/nar/gkm006
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发表时间：
2007
影响因子：
14.9
通讯作者：
Tomkinson AE
中科院分区：
文献类型：
--
作者：
Vijayakumar S;Chapados BR;Schmidt KH;Kolodner RD;Tainer JA;Tomkinson AE
有令人信服的证据表明，增殖细胞核抗原（PCNA）是一种DNA滑动钳，在真核生物DNA复制过程中协调冈崎片段的加工和连接。然而，对功能性PCNA与连接酶I相互作用的详细机制了解尚不完整。在这里，我们展示了酵母PCNA与包含Cdc9的保守的PCNA相互作用基序的肽的共晶结构，酵母DNA连接酶i。Cdc9肽与PCNA的域间连接器环（IDCL）和c端附近的残基接触。互补的突变和生化结果表明，无论是在没有DNA的情况下，还是在PCNA与DNA拓扑相连的情况下，这两个相互作用界面都是复杂形成所必需的。与功能同源的人蛋白类似，酵母RFC与Cdc9 DNA连接酶相互作用并抑制Cdc9 DNA连接酶，而添加PCNA可减轻RFC的抑制作用。本研究表明，PCNA克服rfc介导的Cdc9抑制的能力依赖于PCNA的IDCL和c端相互作用界面。总之，这些结果证明了在PCNA和Cdc9的c端结构域之间形成的β-拉链结构的功能意义，并揭示了FEN-1和Cdc9与两个PCNA界面相互作用的差异，这可能有助于这些酶的协调，顺序作用。
There is compelling evidence that proliferating cell nuclear antigen (PCNA), a DNA sliding clamp, co-ordinates the processing and joining of Okazaki fragments during eukaryotic DNA replication. However, a detailed mechanistic understanding of functional PCNA:ligase I interactions has been incomplete. Here we present the co-crystal structure of yeast PCNA with a peptide encompassing the conserved PCNA interaction motif of Cdc9, yeast DNA ligase I. The Cdc9 peptide contacts both the inter-domain connector loop (IDCL) and residues near the C-terminus of PCNA. Complementary mutational and biochemical results demonstrate that these two interaction interfaces are required for complex formation both in the absence of DNA and when PCNA is topologically linked to DNA. Similar to the functionally homologous human proteins, yeast RFC interacts with and inhibits Cdc9 DNA ligase whereas the addition of PCNA alleviates inhibition by RFC. Here we show that the ability of PCNA to overcome RFC-mediated inhibition of Cdc9 is dependent upon both the IDCL and the C-terminal interaction interfaces of PCNA. Together these results demonstrate the functional significance of the β-zipper structure formed between the C-terminal domain of PCNA and Cdc9 and reveal differences in the interactions of FEN-1 and Cdc9 with the two PCNA interfaces that may contribute to the co-ordinated, sequential action of these enzymes.