Multimerization domains are associated with apparent strand exchange activity in BLM and WRN DNA helicases.
Multimerization domains are associated with apparent strand exchange activity in BLM and WRN DNA helicases.
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DOI：
10.1016/j.dnarep.2014.07.015
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发表时间：
2014-10
期刊：
影响因子：
3.8
通讯作者：
Brill SJ
中科院分区：
文献类型：
--
作者：
Chen CF;Brill SJ
BLM和WRN是RecQ DNA解旋酶家族的成员，其作用是抑制基因组不稳定性和癌症易感性。除了RecQ解旋酶结构域之外，这些蛋白质中的每一个都含有不完全表征的约500个氨基酸（aa）的N-末端结构域。在此之前，我们发现，N-末端的SGS 1，酵母直系同源的BLM，包含一个生理上重要的200个氨基酸的结构域（SGS 1103 -322），显示单链DNA（ssDNA）结合，链退火（SA），并在体外明显的链交换（SE）活动。在这里，我们使用了一种遗传分析，以寻找异源蛋白，可以在体内功能上取代这个域的Sgs 1。与Rad 59相反，寡聚Rad 52蛋白提供体内互补，表明多聚化在功能上是重要的。还鉴定了WRN的N端结构域，其可以在酵母中取代Sgs 1103 -322。该结构域WRN 235 -526包含已知的卷曲螺旋，并显示与Sgs 1103 -322相同的SA和SE活性。发现WRN 235 -526的卷曲螺旋结构域是其体内活性和体外SE活性所需的。基于该结果，在Sgs 1103 -322中识别出潜在的卷曲螺旋。这25个氨基酸的区域同样是必需的野生型Sgs 1在体内的活性，并可替换的异源卷曲螺旋。两者合计，结果表明，卷曲螺旋和紧密相连的表观SE活性是BLM和WRN DNA解旋酶的保守特征。
BLM and WRN are members of the RecQ family of DNA helicases that act to suppress genome instability and cancer predisposition. In addition to a RecQ helicase domain, each of these proteins contains an N-terminal domain of approximately 500 amino acids (aa) that is incompletely characterized. Previously, we showed that the N-terminus of Sgs1, the yeast ortholog of BLM, contains a physiologically important 200 aa domain (Sgs1103–322) that displays single-stranded DNA (ssDNA) binding, strand annealing (SA), and apparent strand-exchange (SE) activities in vitro. Here we used a genetic assay to search for heterologous proteins that could functionally replace this domain of Sgs1 in vivo. In contrast to Rad59, the oligomeric Rad52 protein provided in vivo complementation, suggesting that multimerization is functionally important. An N-terminal domain of WRN was also identified that could replace Sgs1103–322 in yeast. This domain, WRN235–526, contains a known coiled coil and displays the same SA and SE activities as Sgs1103–322. The coiled coil domain of WRN235–526 was found to be required for both its in vivo activity and its in vitro SE activity. Based on this result, a potential coiled coil was identified within Sgs1103–322. This 25 amino acid region was similarly essential for wt Sgs1 activity in vivo and was replaceable by a heterologous coiled coil. Taken together, the results indicate that a coiled coil and a closely-linked apparent SE activity are conserved features of the BLM and WRN DNA helicases.
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