Reinberg D, Zipursky S L, Weisbeek P, Brown D, Hurwitz J
J Biol Chem. 1983 Jan 10;258(1):529-37.
Recombinant RF (replicate form) I DNAs containing the bacteriophage phi X174 gene A protein-recognition sequence are cleaved by the phi X A protein yielding a phi X RF II X A protein complex (Zipursky, S.L., Reinberg, D., and Hurwitz, J. (1980) Proc. Natl. Acad. Sci. U.S.A. 77, 5182-5186). Such complexes support DNA synthesis in both RF I leads to SS(c) and RF I leads to RF I phi X DNA replication reactions in vitro. Two phi X A protein-recognition sequences were inserted into plasmid pBR322. Both sequences were contiguous with the same strand of the vector DNA and separated by 667 and 4275 base pairs. This recombinant plasmid (G27-4) was cleaved by the phi X A protein at either insert and both inserts support the initiation of RF leads to SS(c) DNA synthesis. This was verified by the finding that replication products were circular molecules of 667 and 4275 nucleotides. This finding is in keeping with the multifunctional activities associated with the phi X A protein; these include the site-specific nicking of RF I DNA which initiates DNA synthesis and site-specific termination resulting in the circularization of the displaced DNA strand. The phi X A protein and the Escherichia coli rep and SSb proteins catalyze the unwinding of phi X RF I DNA in vitro (Scott, J.F., Eisenberg, S., Bertsch, L.L., and Kornberg, A. (1977) Proc. Natl. Acad. Sci. U.S.A. 74, 193-197). Recombinant plasmid G27-4 RF I DNA was also unwound in vitro by this enzyme system; in this case, both circular and linear single-stranded DNA molecules of 667 and 4275 nucleotides, as well as full length circular single-stranded DNA were formed. Full length linear DNA was not detected. The two single-stranded circular DNA products formed as leading strands in RF leads to SS(c) reaction mixtures containing G27-4 RF I DNA differed in their ability to support lagging strand DNA synthesis. It was shown that the large single-stranded circular product included DNA sequences homologous to a replication factor Y effector sequence required for RF leads to RF and SS(c) leads to RF replication (Zipursky, S.L., and Marians, K.J. (1980) Proc. Natl. Acad. Sci. U.S.A. 77, 6521-6525). The 4275-nucleotide, but not the 667-nucleotide, single-stranded circular DNA product was converted to a duplex structure.
含有噬菌体φX174基因A蛋白识别序列的重组复制型（RF）I DNA被φX A蛋白切割，产生φX RF II - X A蛋白复合物（齐普斯基，S.L.，赖因贝格，D.，和赫维茨，J.（1980年）《美国国家科学院院刊》77，5182 - 5186）。此类复合物在体外支持RF I→单链（c）和RF I→RF I φX DNA复制反应中的DNA合成。将两个φX A蛋白识别序列插入质粒pBR322。两个序列都与载体DNA的同一条链相邻，间隔分别为667和4275个碱基对。这种重组质粒（G27 - 4）在任一插入位点被φX A蛋白切割，且两个插入序列都支持RF→单链（c）DNA合成的起始。复制产物是667和4275个核苷酸的环状分子，这一发现证实了上述情况。这一发现与φX A蛋白的多功能活性相符；这些活性包括引发DNA合成的RF I DNA位点特异性切口以及导致被置换DNA链环化的位点特异性终止。φX A蛋白与大肠杆菌rep和SSb蛋白在体外催化φX RF I DNA的解旋（斯科特，J.F.，艾森伯格，S.，伯奇，L.L.，和科恩伯格，A.（1977年）《美国国家科学院院刊》74，193 - 197）。重组质粒G27 - 4 RF I DNA在体外也被该酶系统解旋；在这种情况下，形成了667和4275个核苷酸的环状和线性单链DNA分子，以及全长环状单链DNA。未检测到全长线性DNA。在含有G27 - 4 RF I DNA的RF→单链（c）反应混合物中作为前导链形成的两种单链环状DNA产物在支持滞后链DNA合成的能力上有所不同。结果表明，较大的单链环状产物包含与RF→RF和单链（c）→RF复制所需的复制因子Y效应序列同源的DNA序列（齐普斯基，S.L.，和马里安斯，K.J.（1980年）《美国国家科学院院刊》77，6521 - 6525）。4275个核苷酸的单链环状DNA产物而非667个核苷酸的单链环状DNA产物被转化为双链结构。
