A putative mobile genetic element carrying a novel type IIF restriction-modification system (PluTI).
A putative mobile genetic element carrying a novel type IIF restriction-modification system (PluTI).
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DOI：
10.1093/nar/gkp1221
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发表时间：
2010-05
影响因子：
14.9
通讯作者：
Kobayashi I
中科院分区：
文献类型：
--
作者：
Khan F;Furuta Y;Kawai M;Kaminska KH;Ishikawa K;Bujnicki JM;Kobayashi I
利用基因组比较和基因组背景分析，在发光光杆菌（Photorhabdus luminescens）基因组中发现了一个可能的移动的元件。该元件由位于末端区域的16-bp的同向重复序列组成，其与插入序列（IS）的一部分、DNA甲基转移酶基因同源物、两个未知功能的基因和一个开放阅读框（ORF）（plu 0599）编码的蛋白质与任何已知蛋白质没有可检测的序列相似性。当在无细胞表达系统中表达时，ORF（plu 0599）产物显示DNA内切酶活性。随后，将该蛋白命名为R.PluTI，在体内表达，纯化并发现其为识别5′-GGCGC/C-3′（/表示切割位置）的新型IIF限制性内切酶。R.PlutI在两个位点处切割双位点超螺旋底物比单位点超螺旋底物更快。plu 0600编码的修饰酶同源物，命名为M.PluTI，在大肠杆菌中表达，并显示出在体外保护DNA免受R.PluTI切割，并在体内抑制R.PluTI表达的致死作用。这些结果表明，它们构成了一个限制-修饰系统，存在于假定的移动的元素上。因此，我们的方法允许检测以前未表征的DNA相互作用蛋白质家族。
Genome comparison and genome context analysis were used to find a putative mobile element in the genome of Photorhabdus luminescens, an entomopathogenic bacterium. The element is composed of 16-bp direct repeats in the terminal regions, which are identical to a part of insertion sequences (ISs), a DNA methyltransferase gene homolog, two genes of unknown functions and an open reading frame (ORF) (plu0599) encoding a protein with no detectable sequence similarity to any known protein. The ORF (plu0599) product showed DNA endonuclease activity, when expressed in a cell-free expression system. Subsequently, the protein, named R.PluTI, was expressed in vivo, purified and found to be a novel type IIF restriction enzyme that recognizes 5′-GGCGC/C-3′ (/ indicates position of cleavage). R.PluTI cleaves a two-site supercoiled substrate at both the sites faster than a one-site supercoiled substrate. The modification enzyme homolog encoded by plu0600, named M.PluTI, was expressed in Escherichia coli and shown to protect DNA from R.PluTI cleavage in vitro, and to suppress the lethal effects of R.PluTI expression in vivo. These results suggested that they constitute a restriction–modification system, present on the putative mobile element. Our approach thus allowed detection of a previously uncharacterized family of DNA-interacting proteins.
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影响因子：
14.9
作者：
Ishikawa, Ken;Watanabe, Miki;Kuroita, Toshihiro;Uchiyama, Ikuo;Bujnicki, Janusz M;Kawakami, Bunsei;Tanokura, Masaru;Kobayashi, Ichizo
通讯作者：
Kobayashi, Ichizo
影响因子：
14.9
作者：
GINGERAS, TR;MILAZZO, JP;ROBERTS, RJ
通讯作者：
ROBERTS, RJ
影响因子：
14.9
作者：
Grazulis, S;Deibert, M;Siksnys, V
通讯作者：
Siksnys, V
DOI：
10.1038/79032
发表时间：
2000-09-01
期刊：
NATURE STRUCTURAL BIOLOGY
影响因子：
--
作者：
Deibert, M;Grazulis, S;Huber, R
通讯作者：
Huber, R
影响因子：
5.6
作者：
Embleton, ML;Siksnys, V;Halford, SE
通讯作者：
Halford, SE