Sequence and analysis of the replication region of the Staphylococcus xylosus plasmid pSX267
Sequence and analysis of the replication region of the Staphylococcus xylosus plasmid pSX267
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DOI：
10.1016/s0378-1119(96)00526-4
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发表时间：
1996-12-05
期刊：
影响因子：
3.5
通讯作者：
Bruckner, R
中科院分区：
文献类型：
--
作者：
Gering, M;Gotz, F;Bruckner, R
木质葡萄球菌DSM 20267的29.5kb质粒pSX267在1.8kb的片段上被分离出来，这是在葡萄球菌中自主复制所需的区域。序列分析得到两个开放阅读框REPA和ORF2，分别编码37.2 kDa和13.2 kDa的蛋白质。推测的氨基酸序列与粪肠球菌PAD1复制启动蛋白相似，与粪肠球菌pCF10和瑞士乳杆菌pLJ1编码的两种未知功能蛋白相似，与小鼠干扰素反应元件结合因子1相似。pSX267的REPA基因是复制所必需的，提示它编码pSX267的复制启动蛋白。在REPA中引入移码突变或在其3‘端缺失26个密码子会导致非复制性质粒。去除REPA表达所需的序列也取消了复制。与反式REPA的互补实验确定了pSX267在REPA编码区的ORI。位于REPA下游的第二个orf可以在不影响质粒复制的情况下被删除。由于其推导的氨基酸序列与pC194/pUB110-型复制蛋白相似，因此它似乎是pC194/pUB110家族的一个无功能的滚环性质粒残基。定义了pSX267的最小复制子。到目前为止包含1255个碱基对。除REPA外，不需要其他由质粒编码的基因来介导葡萄球菌的复制。
The region of the 29.5-kb plasmid pSX267 from Staphylococcus xylosus DSM 20267 that is required for autonomous replication in staphylococci was isolated on a 1.8-kb DNA fragment. The sequence analysis of the fragment yielded two open reading frames, repA and orf2, encoding proteins of 37.2 and 13.2 kDa, respectively. The deduced amino acid sequence of repA showed similarity to the replication initiator protein of plasmid pAD1 from Enterococcus faecalis, to two proteins of unknown function encoded by the E. faecalis plasmid pCF10 and the Lactobacillus helveticus plasmid pLJ1, and surprisingly to the mouse interferon-response element binding factor 1. The repA gene of pSX267 is indispensable for replication suggesting that it encodes the replication initiator protein of pSX267. Introduction of a frameshift mutation into repA or deletion of 26 codons at its 3'-end resulted in nonreplicative plasmids. Removal of sequences required for repA expression also abolished replication. Complementation experiments with repA in trans identified the ori of pSX267 within the repA coding region. The second orf, which is located downstream of repA, could be deleted without affecting plasmid replication. It seems to be a nonfunctional remainder of a rolling circle plasmid of the pC194/pUB110 family, since its deduced amino acid sequence resembles the pC194/pUB110-type replication proteins. The minimal replicon of pSX267 defined. so far comprises 1255 bp. Besides repA, no other plasmid-encoded gene is required to mediate replication in staphylococci.