Ballester S, Lopez P, Espinosa M, Alonso J C, Lacks S A
Centro de Investigaciones Biológicas, Consejo Superior de Investigaciones Científicas, Madrid, Spain.
J Bacteriol. 1989 May;171(5):2271-7. doi: 10.1128/jb.171.5.2271-2277.1989.
The hybrid plasmid pJS37 is composed of the streptococcal plasmid pLS1, which confers tetracycline resistance, and the staphylococcal plasmid pC194, which confers chloramphenicol resistance. When gram-positive bacteria containing pJS37 were grown in the presence of chloramphenicol, four different deleted derivatives accumulated. The deletions in the plasmid enhanced resistance to chloramphenicol by placing the cat gene of pC194 near promoters of pLS1. All four deletions shared a common endpoint that corresponded to the putative target site for DNA strand nicking by the pC194 replication protein, RepH. At the other, variable endpoint, the DNA sequence was similar to the putative RepH target sequence. Alteration of the RepH protein, by in vitro modification of the gene encoding it, eliminated this class of deletions. By extending a previously proposed model for the generation of a different but related class of deletions (B. Michel and S.D. Ehrlich, EMBO J. 5:3691-3696, 1986), a comprehensive model that could generate both classes of deletions is suggested. It proposes that a nicking-closing activity of the plasmid replication protein at its normal target site and, aberrantly, at sites with similar sequence can generate deletions either proximal or distal to the aberrant site during rolling-circle replication of the plasmid.
杂种质粒pJS37由赋予四环素抗性的链球菌质粒pLS1和赋予氯霉素抗性的葡萄球菌质粒pC194组成。当含有pJS37的革兰氏阳性菌在氯霉素存在下生长时，积累了四种不同的缺失衍生物。质粒中的缺失通过将pC194的cat基因置于pLS1的启动子附近而增强了对氯霉素的抗性。所有四种缺失都有一个共同的终点，该终点对应于pC194复制蛋白RepH对DNA链进行切口的假定靶位点。在另一个可变终点处，DNA序列与假定的RepH靶序列相似。通过对编码RepH蛋白的基因进行体外修饰来改变RepH蛋白，消除了这类缺失。通过扩展先前提出的关于产生另一类不同但相关缺失的模型（B. Michel和S.D. Ehrlich，《欧洲分子生物学组织杂志》5:3691 - 3696，1986年），提出了一个能够产生这两类缺失的综合模型。该模型提出，质粒复制蛋白在其正常靶位点以及异常地在具有相似序列的位点处的切口 - 封闭活性，在质粒滚环复制过程中可在异常位点的近端或远端产生缺失。
