Tosini F, Venanzi S, Boschi A, Battaglia P A
Laboratorio di Biologia Cellulare, Reparto di Genetica Molecolare, Istituto Superiore di Sanità, Rome, Italy.
Mol Gen Genet. 1998 May;258(4):404-11. doi: 10.1007/s004380050748.
The product of the uvp1 gene of the R46 plasmid, a member of the DNA invertase-resolvase family, was studied to characterize its recombination activity on the R46 plasmid. The purified Uvp1 protein specifically binds to a 256-bp DNA fragment located immediately upstream of the uvp1 gene itself, and overlapping the 5'-conserved segment (5'-CS) of the R46 integron In1. We identified on this fragment a putative resolution (res) site. Using an in vitro assay, we demonstrated the ability of the protein to resolve a synthetic cointegrate containing a direct repeat of the res site. In vivo, we obtained cointegrate resolution in Uvp1-expressing recA- cells. Sites I and II, subsites of the putative res site, lie within the outer boundary of the integron 5'-CS which is common to all the known integrons. Furthermore, a 69-bp DNA element (containing site I) is required for cointegrate resolution. We propose that this recombination mechanism protects R46 plasmid against unequal distribution following fusion with either identical or different integron-bearing plasmids. Moreover, Uvp1 might have a role in generating gene cassette diversity between the two conserved segments of the integron.
R46质粒是DNA转化酶-解离酶家族的成员之一,对其uvp1基因的产物进行了研究,以表征其在R46质粒上的重组活性。纯化的Uvp1蛋白特异性结合于uvp1基因自身上游紧邻的一个256bp DNA片段,该片段与R46整合子In1的5'-保守区段(5'-CS)重叠。我们在该片段上鉴定出一个假定的解离(res)位点。通过体外试验,我们证明了该蛋白能够解离一个含有res位点直接重复序列的合成共整合体。在体内,我们在表达Uvp1的recA-细胞中实现了共整合体的解离。假定res位点的亚位点I和II位于所有已知整合子共有的整合子5'-CS的外边界内。此外,共整合体解离需要一个69bp的DNA元件(包含位点I)。我们提出,这种重组机制可保护R46质粒在与携带相同或不同整合子的质粒融合后不会出现不均等分布。此外,Uvp1可能在整合子的两个保守区段之间产生基因盒多样性方面发挥作用。