ENZYMATIC REPLICATION OF THE ORIGIN OF THE ESCHERICHIA-COLI CHROMOSOME
ENZYMATIC REPLICATION OF THE ORIGIN OF THE ESCHERICHIA-COLI CHROMOSOME
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DOI：
10.1073/pnas.78.12.7370
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发表时间：
1981-01-01
期刊：
影响因子：
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通讯作者：
KORNBERG, A
中科院分区：
文献类型：
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作者：
FULLER, RS;KAGUNI, JM;KORNBERG, A
研究了复制带有大肠杆菌染色体起源(ORIC)的质粒的酶系统的相关生理特性。该系统完全依赖于低水平的外源提供的超螺旋ORIC质粒，只使用那些包含.apprx的完整ORIC区域的质粒。245个碱基对，并在ORIC序列内或附近启动复制并双向进行。在这个系统中，复制在5分钟滞后后30-40分钟线性进行，以产生比输入DNA高达40%的产量，依赖于RNA聚合酶和旋转酶，如利福平和奈利地索完全抑制所示，依赖于复制蛋白(例如，dna B蛋白和单链DNA结合蛋白)，如通过特异性抗体抑制判断的，并且独立于蛋白质合成而运行。该系统中的复制还依赖于DNAA的活性，从两个DNAA温度敏感突变株的酶组分的失活和从克隆在多拷贝质粒中的含有DNAA基因的菌株的一组分的互补(互补活性的超产15倍)可以看出这一点。控制染色体周期启动的因素的解析和分析应该通过这个酶系统变得容易。
The physiologically relevant features of an enzyme system that replicates plasmids bearing the origin of the E. coli chromosome (oriC) were studied. The system depends completely on low levels of exogenously furnished supercoiled oriC plasmids, uses only those plasmids that contain the intact oriC region of .apprx. 245 base pairs, and initiates replication within or near the oriC sequence and proceeds bidirectionally. Replication in this system proceeds linearly, after a 5-min lag, for 30-40 min to produce as much as a 40% increase over the input DNA, depends on RNA polymerase and gyrase as indicated by total inhibition by rifampicin and nalidixate, depends on replication proteins (e.g., dnaB protein and single-stranded DNA binding protein) as judged by specific antibody inhibitions and operates independently from protein synthesis. Replication in this system also depended on dnaA activity, as suggested by the inactivity of enzyme fractions from each of 2 dnaA temperature-sensitive mutant strains, and complementation (with a 15-fold overproduction of complementing activity) by a fraction from a strain containing the dnaA gene cloned in a multicopy plasmid. Resolution and analysis of factors that control the initiation of a chromosome cycle should become accessible through this enzyme system.