Imber R, Low R L, Ray D S
Proc Natl Acad Sci U S A. 1983 Dec;80(23):7132-6. doi: 10.1073/pnas.80.23.7132.
A primosome assembly site for F plasmid DNA replication has been identified. This site, which we term rriA (F), is localized to one strand of a 385-base-pair Sau3A restriction fragment very close to ori 2 and within the 2.25-kilobase DNA sequence required for replication and incompatibility of the entire F plasmid. rriA (F) was isolated by cloning into the deletion phage vector M13 delta Elac. This phage forms very faint plaques due to a deletion of the M13 complementary strand origin but forms large wild-type plaques when DNA single-strand initiation determinants are inserted. The single-stranded viral DNA of the Sau3A F-M13 delta Elac recombinant provides an effector site of dATP hydrolysis by the primosomal protein n'. It also provides an assembly site for the Escherichia coli primosome protein complex that directs the in vitro conversion of the single-stranded DNA to a double-stranded form by the same mechanism as that used by phi X174. Homologies of the nucleotide sequence between this F DNA sequence and the previously identified primosome assembly sites in phi X174 phage DNA and in ColE1 plasmid DNA (rriA and rriB) have been found. The sequences 5' G-T-G-A-G-C-G 3' and 5' G-N-G-G-A-A-G-C 3' or variations of these sequences occur from two to five times within each assembly locus. In addition, two distinct 15-base-pair sequences in rriA (F) are perfectly homologous to corresponding sequences in rriA (ColE1).
已鉴定出F质粒DNA复制的引发体组装位点。我们将此位点称为rriA（F），它位于一个385碱基对的Sau3A限制片段的一条链上，非常靠近ori 2，且在整个F质粒复制和不相容性所需的2.25千碱基DNA序列内。rriA（F）是通过克隆到缺失噬菌体载体M13 delta Elac中分离得到的。由于M13互补链起始位点的缺失，该噬菌体形成非常微弱的噬菌斑，但当插入DNA单链起始决定簇时会形成大的野生型噬菌斑。Sau3A F-M13 delta Elac重组体的单链病毒DNA提供了引发体蛋白n'水解dATP的效应位点。它还为大肠杆菌引发体蛋白复合物提供了一个组装位点，该复合物通过与phi X174相同的机制指导体外将单链DNA转化为双链形式。已发现该F DNA序列与先前在phi X174噬菌体DNA和ColE1质粒DNA中鉴定的引发体组装位点（rriA和rriB）之间的核苷酸序列同源性。序列5' G-T-G-A-G-C-G 3'和5' G-N-G-G-A-A-G-C 3'或这些序列的变体在每个组装位点内出现两到五次。此外，rriA（F）中两个不同的15碱基对序列与rriA（ColE1）中的相应序列完全同源。
