The Bacteroides sp. 3_1_23 Pif1 protein is a multifunctional helicase.
The Bacteroides sp. 3_1_23 Pif1 protein is a multifunctional helicase.
复制标题
拟杆菌属。
DOI：
10.1093/nar/gkv916
复制
发表时间：
2015-10-15
影响因子：
14.9
通讯作者：
Xi XG
中科院分区：
文献类型：
--
作者：
Liu NN;Duan XL;Ai X;Yang YT;Li M;Dou SX;Rety S;Deprez E;Xi XG
ScPif1 DNA解旋酶是从细菌到人类保守的5′- 3′解旋酶超家族的典型成员，在维持基因组稳态中发挥多种作用。虽然对真核Pif1解旋酶（包括酵母和人类Pif1蛋白）进行了许多研究，但原核Pif1解旋酶的潜在功能和生化特性在很大程度上仍然未知。本文报道了拟杆菌3_1_23 （Bacteroides sp. 3_1_23, BsPif1）中Pif1解旋酶的表达、纯化和生化分析。BsPif1与大量DNA底物结合，特别是有效地解旋具有5 '悬垂，叉状底物，d -环和flap样底物的部分双工DNA，这表明BsPif1可能在停滞的DNA复制分叉处起作用并促进冈崎片段成熟。与真核生物同源物一样，BsPif1可以分解r环结构并解绕DNA-RNA杂交体。此外，BsPif1有效地展开g -四联体并破坏核蛋白复合物。总之，这些结果强调原核Pif1解旋酶可能解决DNA交易过程中出现的常见问题。有趣的是，我们发现BsPif1与酵母的Pif1不同，但在底物特异性、解旋酶活性和作用方式方面更像人类的Pif1。这些发现在原核生物Pif1解旋酶在体内可能的功能的背景下进行了讨论。
ScPif1 DNA helicase is the prototypical member of a 5′-to-3′ helicase superfamily conserved from bacteria to human and plays various roles in the maintenance of genomic homeostasis. While many studies have been performed with eukaryotic Pif1 helicases, including yeast and human Pif1 proteins, the potential functions and biochemical properties of prokaryotic Pif1 helicases remain largely unknown. Here, we report the expression, purification and biochemical analysis of Pif1 helicase from Bacteroides sp. 3_1_23 (BsPif1). BsPif1 binds to a large panel of DNA substrates and, in particular, efficiently unwinds partial duplex DNAs with 5′-overhang, fork-like substrates, D-loop and flap-like substrates, suggesting that BsPif1 may act at stalled DNA replication forks and enhance Okazaki fragment maturation. Like its eukaryotic homologues, BsPif1 resolves R-loop structures and unwinds DNA–RNA hybrids. Furthermore, BsPif1 efficiently unfolds G-quadruplexes and disrupts nucleoprotein complexes. Altogether, these results highlight that prokaryotic Pif1 helicases may resolve common issues that arise during DNA transactions. Interestingly, we found that BsPif1 is different from yeast Pif1, but resembles more human Pif1 with regard to substrate specificity, helicase activity and mode of action. These findings are discussed in the context of the possible functions of prokaryotic Pif1 helicases in vivo.