Cho Il-Taeg, Kim Do-Hyung, Kang Young-Hoon, Lee Chul-Hwan, Amangyelid Tamir, Nguyen Tuan Anh, Hurwitz Jerard, Seo Yeon-Soo
Center for DNA Replication and Genome Instability, Department of Biological Sciences, Korea Advanced Institute of Science and Technology, Daejeon 305-701, Korea.
J Biol Chem. 2009 Apr 17;284(16):10387-99. doi: 10.1074/jbc.M808893200. Epub 2009 Feb 9.
Flap endonuclease 1 (FEN1) is the enzyme responsible for specifically removing the flap structure produced during DNA replication, repair, and recombination. Here we report that the human replication factor C (RFC) complex stimulates the nuclease activity of human FEN1 in an ATP-independent manner. Although proliferating cell nuclear antigen is also known to stimulate FEN1, less RFC was required for comparable FEN1 stimulation. Kinetic analyses indicate that the mechanism by which RFC stimulates FEN1 is distinct from that by proliferating cell nuclear antigen. Heat-denatured RFC or its subunit retained, fully or partially, the ability to stimulate FEN1. Via systematic deletion analyses, we have defined three specific regions of RFC4 capable of stimulating FEN1. The region of RFC4 with the highest activity spans amino acids 170-194 and contains RFC box VII. Four amino acid residues (i.e. Tyr-182, Glu-188, Pro-189, and Ser-192) are especially important for FEN1 stimulatory activity. Thus, RFC, via several stimulatory motifs per molecule, potently activates FEN1. This function makes RFC a critical partner with FEN1 for the processing of eukaryotic Okazaki fragments.
瓣状核酸内切酶1（FEN1）是一种负责特异性去除DNA复制、修复和重组过程中产生的瓣状结构的酶。在此我们报告，人类复制因子C（RFC）复合体以不依赖ATP的方式刺激人类FEN1的核酸酶活性。尽管已知增殖细胞核抗原也能刺激FEN1，但在产生相当的FEN1刺激效果时所需的RFC较少。动力学分析表明，RFC刺激FEN1的机制与增殖细胞核抗原不同。热变性的RFC或其亚基完全或部分保留了刺激FEN1的能力。通过系统的缺失分析，我们确定了RFC4中能够刺激FEN1的三个特定区域。RFC4中活性最高的区域跨度为氨基酸170 - 194，包含RFC框VII。四个氨基酸残基（即Tyr - 182、Glu - 188、Pro - 189和Ser - 192）对FEN1刺激活性尤为重要。因此，RFC通过每个分子中的几个刺激基序有效地激活FEN1。这一功能使RFC成为FEN1处理真核冈崎片段的关键伙伴。
