Phage ORF family recombinases: conservation of activities and involvement of the central channel in DNA binding.
Phage ORF family recombinases: conservation of activities and involvement of the central channel in DNA binding.
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DOI：
10.1371/journal.pone.0102454
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发表时间：
2014
期刊：
影响因子：
3.7
通讯作者：
Sharples GJ
中科院分区：
文献类型：
--
作者：
Curtis FA;Malay AD;Trotter AJ;Wilson LA;Barradell-Black MM;Bowers LY;Reed P;Hillyar CR;Yeo RP;Sanderson JM;Heddle JG;Sharples GJ
遗传和生化证据表明，λ Orf是一种重组介质，可促进细菌RecA或噬菌体Redβ重组酶在SSB蛋白结合的单链DNA （ssDNA）上成核。我们已经确定了一个多样化的Orf蛋白家族，其中包括涉及DNA碱基翻转的代表和融合到HNH内切酶结构域的代表。为了确认与Orf家族的功能关系，从大肠杆菌隐前噬菌体DLP12中纯化并表征了一个远亲同源物YbcN。和它的λ亲戚一样，YbcN更倾向于结合ssDNA而不是双链。Orf和YbcN都没有表现出对含有错配或1-3个核苷酸凸起的双链DNA的显著偏好。YbcN也与大肠杆菌SSB结合，尽管与Orf不同，它未能与缺乏协调异种蛋白-蛋白相互作用的柔性c端尾的SSB突变体结合。Orf家族中位于λ Orf晶体结构中心空腔两侧的保守残基被靶向诱变以帮助确定DNA结合模式。这些突变蛋白中的一些在DNA结合上显示出明显的缺陷，这与中心孔对底物识别很重要一致。orf样蛋白的广泛保存突出了在羔羊样噬菌体重组过程中靶向SSB包被的ssDNA的重要性。
Genetic and biochemical evidence suggests that λ Orf is a recombination mediator, promoting nucleation of either bacterial RecA or phage Redβ recombinases onto single-stranded DNA (ssDNA) bound by SSB protein. We have identified a diverse family of Orf proteins that includes representatives implicated in DNA base flipping and those fused to an HNH endonuclease domain. To confirm a functional relationship with the Orf family, a distantly-related homolog, YbcN, from Escherichia coli cryptic prophage DLP12 was purified and characterized. As with its λ relative, YbcN showed a preference for binding ssDNA over duplex. Neither Orf nor YbcN displayed a significant preference for duplex DNA containing mismatches or 1-3 nucleotide bulges. YbcN also bound E. coli SSB, although unlike Orf, it failed to associate with an SSB mutant lacking the flexible C-terminal tail involved in coordinating heterologous protein-protein interactions. Residues conserved in the Orf family that flank the central cavity in the λ Orf crystal structure were targeted for mutagenesis to help determine the mode of DNA binding. Several of these mutant proteins showed significant defects in DNA binding consistent with the central aperture being important for substrate recognition. The widespread conservation of Orf-like proteins highlights the importance of targeting SSB coated ssDNA during lambdoid phage recombination.
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DOI：
10.1073/pnas.252633399
发表时间：
2002-11-26
影响因子：
11.1
作者：
Kantake, N;Madiraju, MVVM;Kowalczykowski, C
通讯作者：
Kowalczykowski, C
DOI：
10.1073/pnas.121164898
发表时间：
2001-06-05
影响因子：
11.1
作者：
Ellis, HM;Yu, DG;Court, DL
通讯作者：
Court, DL
影响因子：
56.9
作者：
Courcelle, J;Donaldson, JR;Courcelle, CT
通讯作者：
Courcelle, CT
影响因子：
3.3
作者：
Donaldson, JR;Courcelle, CT;Courcelle, J
通讯作者：
Courcelle, J
影响因子：
14.9
作者：
Finn RD;Mistry J;Tate J;Coggill P;Heger A;Pollington JE;Gavin OL;Gunasekaran P;Ceric G;Forslund K;Holm L;Sonnhammer EL;Eddy SR;Bateman A
通讯作者：
Bateman A