PDF(Pubmed)
Abstract：
Topoisomerase IV (Topo IV) is the main decatenation enzyme in Escherichia coli; it removes catenation links that are formed during DNA replication. Topo IV binding and cleavage sites were previously identified in the E. coli genome with ChIP-Seq and NorfIP. Here, we used a more sensitive, single-nucleotide resolution Topo-Seq procedure to identify Topo IV cleavage sites (TCSs) genome-wide. We detected thousands of TCSs scattered in the bacterial genome. The determined cleavage motif of Topo IV contained previously known cleavage determinants (-4G/+8C, -2A/+6 T, -1 T/+5A) and additional, not observed previously, positions -7C/+11G and -6C/+10G. TCSs were depleted in the Ter macrodomain except for two exceptionally strong non-canonical cleavage sites located in 33 and 38 bp from the XerC-box of the dif-site. Topo IV cleavage activity was increased in Left and Right macrodomains flanking the Ter macrodomain and was especially high in the 50-60 kb region containing the oriC origin of replication. Topo IV enrichment was also increased downstream of highly active transcription units, indicating that the enzyme is involved in relaxation of transcription-induced positive supercoiling.
摘要：
拓扑异构酶IV（TopoIV）是大肠杆菌中的主要去酶；它消除了DNA复制过程中形成的连接连接。先前用ChIP-Seq和NorfIP在大肠杆菌基因组中鉴定了TopoIV结合和切割位点。这里,我们用了一个更敏感的,单核苷酸分辨率Topo-Seq程序，以识别全基因组的TopoIV切割位点（TCSs）。我们检测到数千个散布在细菌基因组中的TCS。确定的TopoIV的裂解基序包含先前已知的裂解决定簇（-4G/8C，-2A/+6T,-1T/+5A)和附加，以前没有观察到，位置-7C/+11G和-6C/+10G。除了位于dif位点的XerC-box的33和38bp中的两个异常强的非规范切割位点外，TCS在Ter宏结构域中被耗尽。TopoIV切割活性在Ter巨域侧翼的左和右巨域中增加，在包含oriC复制起点的50-60kb区域中特别高。TopoIV富集也增加了高活性转录单位的下游，表明该酶参与转录诱导的正超螺旋的松弛。
