Molecular characterization of functional modules of plasmid pWKS1 of Paracoccus pantotrophus DSM 11072
Molecular characterization of functional modules of plasmid pWKS1 of Paracoccus pantotrophus DSM 11072
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DOI：
10.1099/00221287-148-9-2847
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发表时间：
2002-09-01
期刊：
影响因子：
2.8
通讯作者：
Wlodarczyk, M
中科院分区：
文献类型：
--
作者：
Bartosik, D;Baj, J;Wlodarczyk, M
确定了副球菌 DSM 11072 的小型隐秘质粒 pWKS1（2697 bp）的完整核苷酸序列。该质粒序列的G+C含量为62mol％。分析显示，超过 80% 的质粒基因组被两个 ORF（ORF1 和 ORF2）覆盖，它们分别能够编码 44.1 和 37.8 kDa 的推定肽。突变分析表明ORF2对于质粒复制至关重要。 ORF2的翻译产物与几种θ复制乳球菌质粒的复制蛋白以及革兰氏阴性宿主中质粒的Rep蛋白具有局部同源性。富含 A+T 的区域位于rep 基因上游，包含三个串联重复的 21 bp 长 iteron 样序列，作为复制起点 (oriV)。 ORF1 编码一种假定的动员蛋白，与广泛宿主范围质粒 pBBR1 和革兰氏阳性菌质粒的动员蛋白 (Mob) 相似。带有 pWKS1 MOB 模块（mob 基因和 oriT 序列）的质粒可以在不同大肠杆菌菌株之间以低频率进行转移（通过 IncP 11134 转移装置）。 pWKS1 和 pBBR1 的 MOB 模块在功能上互补。杂交分析表明，在迄今为止鉴定的所有副球菌质粒中，只有质粒pSOV1（6.5 kb）携带与pWKS1相关的序列。质粒pWKS1可以在10种副球菌、根癌农杆菌、豆根瘤菌和球形红杆菌中复制，但不能在大肠杆菌中复制。
The complete nucleotide sequence of the small, cryptic plasmid pWKS1 (2697 bp) of Paracoccus pantotrophus DSM 11072 was determined. The G+C content of the sequence of this plasmid was 62 mol %. Analysis revealed that over 80 % of the plasmid genome was covered by two ORFs, ORF1 and ORF2, which were capable of encoding putative peptides of 44.1 and 37.8 kDa, respectively. Mutational analysis showed that ORF2 was crucial for plasmid replication. The translational product of ORF2 shared local homologies with replication proteins of several theta-replicating lactococcal plasmids, as well as with the Rep proteins of plasmids residing in Gram-negative hosts. An A+T-rich region, located upstream of the rep gene and containing three tandemly repeated 21 bp long iteron-like sequences, served as the origin of replication (oriV). ORF1 encoded a putative mobilization protein with similarities to mobilization proteins (Mob) from the broad-host-range plasmid pBBR1 and plasmids of Gram-positive bacteria. A plasmid bearing the MOB module of pWKS1 (the mob gene and the oriT sequence) could be mobilized for transfer (by IncP 11134 transfer apparatus) at low frequency between different strains of Escherichia coli. MOB modules of pWKS1 and pBBR1 were functionally complementary to each other. Hybridization analysis revealed that only plasmid pSOV1 (6.5 kb), among all of the paracoccal plasmids identified so far, carries sequences related to pWKS1. Plasmid pWKS1 could replicate in 10 species of Paracoccus and in Agrobacterium tumefaciens, Rhizobium leguminosarum and Rhodobacter sphaeroides, but it could not replicate in E. coli.