DNA STRUCTURAL ELEMENTS REQUIRED FOR FEN-1 BINDING
DNA STRUCTURAL ELEMENTS REQUIRED FOR FEN-1 BINDING
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DOI：
10.1074/jbc.270.9.4503
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发表时间：
1995-03-03
影响因子：
4.8
通讯作者：
LIEBER, MR
中科院分区：
文献类型：
--
作者：
HARRINGTON, JJ;LIEBER, MR
在真核细胞中，5 ′-瓣状DNA内切核酸酶和双链DNA 5 ′-外切核酸酶活性存在于称为FEN-1（瓣状内切核酸酶-1和5（5 ′-外切核酸酶））的42-kDa酶内。酿酒酵母YKL 510。与FEN-1一样，这些相关的结构特异性核酸酶识别并切割称为DNA瓣的分支DNA结构及其衍生物，称为假Y结构。为了剖析DNA瓣结构的重要结构组分，我们开发了迁移率变动测定法，我们发现F-形容词链（位于移位的瓣链附近）是FEN-1有效结合和切割瓣结构所必需的。当该链不存在或存在时，但从瓣链手肘凹进，结合效率下降。使用双瓣结构对F-adj链的作用的进一步研究揭示，F-adj链对于提供FEN-1可以在瓣链手肘附近结合的双链模板是必需的，这些结果为理解这种结构特异性核酸酶如何识别各种DNA底物提供了基础。
In eukaryotic cells, a 5'-flap DNA endonuclease and a double-stranded DNA 5'-exonuclease activity reside within a 42-kDa enzyme called FEN-1 (flap endonuclease-1 and 5(five)'-exonuclease), This endo/exonuclease has been shown to be highly homologous to human XP-G, Saccharomyces cerevisiae RAD2, and S. cerevisiae YKL510. Like FEN-1, these related structure-specific nucleases recognize and cleave a branched DNA structure called a DNA flap and its derivative, called a pseudo Y-structure, To dissect the important structural components of the DNA flap structure, we have developed a mobility shift assay, We find that the F-adj strand (located adjacent to the displaced flap strand) is necessary for efficient binding and cleavage of flap structures by FEN-1. When this strand is absent or when it is present, but recessed from the elbow of the flap strand, binding efficiency drops, Further investigation of the role of the F-adj strand using double flap structures reveals that the F-adj strand is necessary to provide a double-stranded template upon which FEN-1 can bind near the elbow of the flap strand, These results provide a basis for understanding how this structure-specific nuclease recognizes a variety of DNA substrates.