Binding of SeqA protein to DNA requires interaction between two or more complexes bound to separate hemimethylated GATC sequences
Binding of SeqA protein to DNA requires interaction between two or more complexes bound to separate hemimethylated GATC sequences
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DOI：
10.1093/emboj/18.8.2304
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发表时间：
1999-04-15
期刊：
影响因子：
11.4
通讯作者：
Austin, S
中科院分区：
文献类型：
--
作者：
Brendler, T;Austin, S
SeqA蛋白在半甲基化GATC腺嘌呤甲基化位点与大肠杆菌染色体（oriC）和P1质粒（P1oriR）复制起点的复制后形式结合。它似乎通过防止过早的重新启动来调节复制。然而，SeqA结合并不限于复制起点：当某些规则适用时，具有半甲基化GATC位点的不同片段可以在体外结合SeqA。最值得注意的是，在结合片段上必须存在多于一个这样的位点。该蛋白质似乎认识到个人的半甲基化位点，但必须经历一个专性的合作与附近的结合蛋白质的稳定结合的相互作用。SeqA在每个半甲基化的GATC序列处以离散的补丁接触两条DNA链。所有四个GATC碱基都是接触的，并且是结合所必需的。虽然识别的序列是对称的，但甲基化链上的足迹总是更宽，这表明结合的蛋白质是不对称定位的，其方向由独特的甲基基团的位置决定。替代间距和相邻位点的相对取向的研究表明，每个位点可以被识别的一个诱导的不对称性的亚基之一，与某些II型限制性内切酶的对称二聚体。
The SeqA protein binds to the post-replicative forms of the origins of replication of the Escherichia coli chromosome (oriC) and the P1 plasmid (P1oriR) at hemimethylated GATC adenine methylation sites. It appears to regulate replication by preventing premature reinitiation. However, SeqA binding is not exclusive to replication origins: different fragments with hemimethylated GATC sites can bind SeqA in vitro when certain rules apply. Most notably, more than one such site must be present on a bound fragment. The protein appears to recognize individual hemimethylated sites, but must undergo an obligate cooperative interaction with a nearby bound protein for stable binding. SeqA contacts both DNA strands in a discrete patch at each hemimethylated GATC sequence. All four GATC bases are contacted and are essential for binding. Although the recognized sequence is symmetrical, the footprint on the methylated strand is always broader, suggesting that the bound protein is positioned asymmetrically with its orientation dictated by the position of the unique methyl group. Studies of alternative spacings and relative orientations of adjacent sites suggest that each site may be recognized by a symmetrical dimer with an induced asymmetry in one of the subunits similar to that seen with certain type II restriction endonucleases.